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Sample records for human apolipoprotein ai

  1. Opposite regulation of human versus mouse apolipoprotein A-I by fibrates in human apolipoprotein A-I transgenic mice.

    PubMed Central

    Berthou, L; Duverger, N; Emmanuel, F; Langouët, S; Auwerx, J; Guillouzo, A; Fruchart, J C; Rubin, E; Denèfle, P; Staels, B; Branellec, D

    1996-01-01

    The regulation of liver apolipoprotein (apo) A-I gene expression by fibrates was studied in human apo A-I transgenic mice containing a human genomic DNA fragment driving apo A-I expression in liver. Treatment with fenofibrate (0.5% wt/wt) for 7 d increased plasma human apo A-I levels up to 750% and HDL-cholesterol levels up to 200% with a shift to larger particles. The increase in human apo A-I plasma levels was time and dose dependent and was already evident after 3 d at the highest dose (0.5% wt/wt) of fenofibrate. In contrast, plasma mouse apo A-I concentration was decreased after fenofibrate in nontransgenic mice. The increase in plasma human apo A-I levels after fenofibrate treatment was associated with a 97% increase in hepatic human apo A-I mRNA, whereas mouse apo A-I mRNA levels decreased to 51%. In nontransgenic mice, a similar down-regulation of hepatic apo A-I mRNA levels was observed. Nuclear run-on experiments demonstrated that the increase in human apo A-I and the decrease in mouse apo A-I gene expression after fenofibrate occurred at the transcriptional level. Since part of the effects of fibrates are mediated through the nuclear receptor PPAR (peroxisome proliferator-activated receptor), the expression of the acyl CoA oxidase (ACO) gene was measured as a control of PPAR activation. Both in transgenic and nontransgenic mice, fenofibrate induced ACO mRNA levels up to sixfold. When transgenic mice were treated with gemfibrozil (0.5% wt/wt) plasma human apo A-I and HDL-cholesterol levels increased 32 and 73%, respectively, above control levels. The weaker effect of this compound on human apo A-I and HDL-cholesterol levels correlated with a less pronounced impact on ACO mRNA levels (a threefold increase) suggesting that the level of induction of human apo A-I gene is related to the PPAR activating potency of the fibrate used. Treatment of human primary hepatocytes with fenofibric acid (500 microM) provoked an 83 and 50% increase in apo A-I secretion and

  2. The apolipoprotein CIII enhancer regulates both extensive histone modification and intergenic transcription of human apolipoprotein AI/CIII/AIV genes but not apolipoprotein AV.

    PubMed

    Li, Ya-Jun; Wei, Yu-Sheng; Fu, Xiang-Hui; Hao, De-Long; Xue, Zheng; Gong, Huan; Zhang, Zhu-Qin; Liu, De-Pei; Liang, Chih-Chuan

    2008-10-17

    The apolipoprotein (apo) AI/CIII/AIV/AV cluster genes are expressed at different levels in the liver and intestine. The apoCIII enhancer, a common regulatory element, regulates the tissue-specific expression of apoAI, apoCIII, and apoAIV but not apoAV. To study this regulation at the chromatin level, the histone modifications and intergenic transcription in the human apoAI/CIII/AIV/AV cluster were investigated in HepG2 and Caco-2 cells and in the livers of transgenic mice carrying the human gene cluster constructs with or without the apoCIII enhancer. We found that both the promoters and the intergenic regions of the apoAI/CIII/AIV genes were hyperacetylated and formed an open subdomain that did not include the apoAV gene. Hepatic and intestinal intergenic transcripts were identified to transcribe bidirectionally with strand preferences along the cluster. The deletion of the apoCIII enhancer influenced both histone modification and intergenic transcription in the apoAI/CIII/AIV gene region. These results demonstrate that the apoCIII enhancer contributes to the maintenance of an active chromatin subdomain of the apoAI/CIII/AIV genes, but not apoAV.

  3. [Separation and purification of human apolipoproteins A-I and C-III by chromatofocusing].

    PubMed

    Cheng, B

    1993-08-01

    Human very low density lipoprotein (VLDL) and high density lipoprotein (HDL) were isolated and purified by a process of combined dextran sulfate precipitation and density gradient ultracentrifugation. Chromatofocusing, which separates protein based on differences in isoelectric point, was used to separate apolipoprotein A-I (apoA-I) and apolipoprotein C-III from human HDL and VLDL, respectively. Discontinuous SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and analytical isoelectric focusing (IEF) were used to study the purity of different fractions. Both purified apoA-I and apoC-III showed single bands on SDS-PAGE at molecular weights of 28183 and 9400 Daltons, respectively. As determined by IEF in the presence of 8 mol/L urea, apoA-I had eight isoforms with pI of 5.66-5.87. The pI's of the three isoproteins of apoC-III (C-III0, C-III1 and C-III2) were 5.06, 4.88 and 4.72, respectively. Chromatofocusing, a new simple technique combining the high resolving power of IEF with the high capacity of ion-exchange column chromatography, is extremely valuable for large-scale purification of the major apolipoproteins of VLDL and HDL.

  4. A complete backbone spectral assignment of human apolipoprotein AI on a 38 kDa preβHDL (Lp1-AI) particle

    SciTech Connect

    Ren, Xuefeng; Yang, Yunhuang; Neville, T.; Hoyt, David W.; Sparks, Daniel L.; Wang, Jianjun

    2007-06-12

    Apolipoprotein A-I (apoAI, 243-residues) is the major protein component of the high-density lipoprotein (HDL) that has been a hot subject of interests because of its anti-atherogenic properties. This important property of apoAI is related to its roles in reverse cholesterol transport pathway. Upon lipid-binding, apoAI undergoes conformational changes from lipid-free to several different HDL-associated states (1). These different conformational states regulate HDL formation, maturation and transportation. Two initial conformational states of apoAI are lipid-free apoAI and apoAI/preβHDL that recruit phospholipids and cholesterol to form HDL particles. In particular, lipid-free apoAI specifically binds to phospholipids to form lipid-poor apoAI, including apoAI/preβ-HDL (~37 kDa). As a unique class of lipid poor HDL, both in vitro and in vivo evidence demonstrates that apoAI/preβ-HDLs are the most effective acceptors specifically for free cholesterol in human plasma and serves as the precursor of HDL particles (2). Here we report a complete backbone spectral assignment of human apoAI/preβHDL. Secondary structure prediction using backbone NMR parameters indicates that apoAI/preβHDL displays a two-domain structure: the N-terminal four helix-bundle domain (residues 1-186) and the C-terminal flexible domain (residues 187-243). A structure of apoAI/preβ-HDL is the first lipid-associated structure of apoAI and is critical for us to understand how apoAI recruits cholesterol to initialize HDL formation. BMRB deposit with accession number: 15093.

  5. Properties of discoidal complexes of human apolipoprotein A-I with phosphatidylcholines containing various fatty acid chains.

    PubMed

    Zorich, N L; Kézdy, K E; Jonas, A

    1987-06-02

    In this study we demonstrate that apolipoprotein A-I determined the common size classes of discoidal particles formed with numerous phosphatidylcholines, and with ether analogs of phosphatidylcholines. We show furthermore, that the nature of the lipids dictates the distribution of particles among the different size classes. These experiments were performed with discoidal complexes containing various phospholipids (phosphatidylcholines with saturated and unsaturated fatty acid chains of different lengths and the ether analog of 1-palmitoyl-2-oleoylphosphatidylcholine), cholesterol, and human apolipoprotein A-I, prepared by the sodium cholate dialysis method, and fractionated by Bio-Gel A-5m gel-filtration chromatography. The complex preparations were analyzed in terms of their average composition, spectral properties of the apolipoprotein, and the dynamic behavior of the lipid domains. Nondenaturing gradient gel electrophoresis was used to analyze the size classes of particles present in the complex preparations. Starting with reaction mixtures containing around 100:1, phospholipid/apolipoprotein A-I molar ratios, complexes were isolated with molar ratios from 40:1 to 100:1. In most complexes apolipoprotein A-I had high levels of alpha-helical structure (65-77% alpha-helix), and tryptophan residues in a nonpolar environment. The lipid domains of complexes exhibited the dynamic behavior expected of the main phospholipid components. In the average size range from 90 to 100 A diameters, discrete particle classes with 80, 87, 102, 108, or 112 A Stokes diameters were observed for all the complexes containing different phospholipids. These discrete, recurring particle sizes are attributed to distinct apolipoprotein A-I conformations and variable lipid content.

  6. Radial-immunodiffusion assay of human apolipoprotein A-I with use of two monoclonal antibodies combined.

    PubMed

    Marcovina, S; Di Cola, G; Catapano, A L

    1986-12-01

    We produced and characterized several monoclonal antibodies directed toward human plasma apolipoprotein A-I. Two of them, A-I-12 and A-I-57, individually precipitated purified or native high-density lipoprotein in agarose gel by double immunodiffusion. Because radial immunodiffusion performed with a single monoclonal antibody gave faint and diffuse rings of precipitation, we developed and optimized working conditions for using these two monoclonal antibodies combined to determine apolipoprotein A-I in human plasma. This combination gave easy-to-measure, clear, sharp rings, and linear and parallel standard curves for HDL3 (the primary standard) and a reference serum (the secondary standard). Moreover, no pretreatment of samples with dissociating agents or detergents is necessary. The assay was complete after overnight incubation, as compared with two to three days when polyclonal antisera were used. Apolipoprotein A-I concentrations as measured in 128 normolipidemic subjects and in 72 patients with various lipid disorders by the radial immunodiffusion technique with monoclonal antibodies (x) compared well (r = 0.882; y = 1.029x-0.036) with those measured by radial immunodiffusion with polyclonal antisera (y).

  7. Expression of the C-terminal domain of human apolipoprotein A-I using a chimeric apolipoprotein.

    PubMed

    Sallee, Daniel E; Horn, James V C; Fuentes, Lukas A; Weers, Paul M M

    2017-09-01

    Human apolipoprotein A-I (apoA-I) is the most abundant protein in high-density lipoprotein, an anti-atherogenic lipid-protein complex responsible for reverse cholesterol transport. The protein is composed of an N-terminal helix bundle domain, and a small C-terminal (CT) domain. To facilitate study of CT-apoA-I, a novel strategy was employed to produce this small domain in a bacterial expression system. A protein construct was designed of insect apolipophorin III (apoLp-III) and residues 179-243 of apoA-I, with a unique methionine residue positioned between the two proteins and an N-terminal His-tag to facilitate purification. The chimera was expressed in E. coli, purified by Ni-affinity chromatography, and cleaved by cyanogen bromide. SDS-PAGE revealed the presence of three proteins with masses of 7 kDa (CT-apoA-I), 18 kDa (apoLp-III), and a minor 26 kDa band of uncleaved chimera. The digest was reloaded on the Ni-affinity column to bind apoLp-III and uncleaved chimera, while CT-apoA-I was washed from the column and collected. Alternatively, CT-apoA-I was isolated from the digest by reversed-phase HPLC. CT-apoA-I was α-helical, highly effective in solubilizing phospholipid vesicles and disaggregating LPS micelles. However, CT-apoA-I was less active compared to full-length apoA-I in protecting lipolyzed low density lipoproteins from aggregating, and disrupting phosphatidylglycerol bilayer vesicles. Thus the novel expression system produced mg quantities of functional CT-apoA-I, facilitating structural and functional studies of this critical domain of apoA-I. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Nucleotide sequence and the encoded amino acids of human apolipoprotein A-I mRNA.

    PubMed Central

    Law, S W; Brewer, H B

    1984-01-01

    The cDNA clones encoding the precursor form of human liver apolipoprotein A-I (apoA-I), preproapoA-I, have been isolated from a cDNA library. A 17-base synthetic oligonucleotide based on residues 108-113 of apoA-I and a 26-base primer-extended, dideoxynucleotide-terminated cDNA were used as hybridization probes to select for recombinant plasmids bearing the apoA-I sequence. The complete nucleic acid sequence of human liver preproapoA-I has been determined by analysis of the cloned cDNA. The sequence is composed of 801 nucleotides encoding 267 amino acid residues. PreproapoA-I contains an 18-amino-acid prepeptide and a 6-amino-acid propeptide connected to the amino terminus of the 243-amino acid mature apoA-I. Southern blotting analysis of chromosomal DNA obtained from peripheral blood indicated the apoA-I gene is contained in a 2.1-kilobase-pair Pst I fragment and there is no gross difference in structural organization between the normal apoA-I gene and the Tangier disease apoA-I gene. Images PMID:6198645

  9. Helical structure and stability in human apolipoprotein A-I by hydrogen exchange and mass spectrometry

    PubMed Central

    Chetty, Palaniappan Sevugan; Mayne, Leland; Lund-Katz, Sissel; Stranz, David; Englander, S. Walter; Phillips, Michael C.

    2009-01-01

    Apolipoprotein A-I (apoA-I) stabilizes anti-atherogenic high density lipoprotein particles (HDL) in the circulation and governs their biogenesis, metabolism, and functional interactions. To decipher these important structure–function relationships, it will be necessary to understand the structure, stability, and plasticity of the apoA-I molecule. Biophysical studies show that lipid-free apoA-I contains a large amount of α-helical structure but the location of this structure and its properties are not established. We used hydrogen-deuterium exchange coupled with a fragmentation-separation method and mass spectrometric analysis to study human lipid-free apoA-I in its physiologically pertinent monomeric form. The acquisition of ≈100 overlapping peptide fragments that redundantly cover the 243-residue apoA-I polypeptide made it possible to define the positions and stabilities of helical segments and to draw inferences about their interactions and dynamic properties. Residues 7–44, 54–65, 70–78, 81–115, and 147–178 form α-helices, accounting for a helical content of 48 ± 3%, in agreement with circular dichroism measurements (49%). At 3 to 5 kcal/mol in free energy of stabilization, the helices are far more stable than could be achieved in isolation, indicating mutually stabilizing helix bundle interactions. However the helical structure is dynamic, unfolding and refolding in seconds, allowing facile apoA-I reorganization during HDL particle formation and remodeling. PMID:19850866

  10. Human Apolipoprotein A-I Natural Variants: Molecular Mechanisms Underlying Amyloidogenic Propensity

    PubMed Central

    Ramella, Nahuel A.; Schinella, Guillermo R.; Ferreira, Sergio T.; Prieto, Eduardo D.; Vela, María E.; Ríos, José Luis

    2012-01-01

    Human apolipoprotein A-I (apoA-I)-derived amyloidosis can present with either wild-type (Wt) protein deposits in atherosclerotic plaques or as a hereditary form in which apoA-I variants deposit causing multiple organ failure. More than 15 single amino acid replacement amyloidogenic apoA-I variants have been described, but the molecular mechanisms involved in amyloid-associated pathology remain largely unknown. Here, we have investigated by fluorescence and biochemical approaches the stabilities and propensities to aggregate of two disease-associated apoA-I variants, apoA-IGly26Arg, associated with polyneuropathy and kidney dysfunction, and apoA-ILys107-0, implicated in amyloidosis in severe atherosclerosis. Results showed that both variants share common structural properties including decreased stability compared to Wt apoA-I and a more flexible structure that gives rise to formation of partially folded states. Interestingly, however, distinct features appear to determine their pathogenic mechanisms. ApoA-ILys107-0 has an increased propensity to aggregate at physiological pH and in a pro-inflammatory microenvironment than Wt apoA-I, whereas apoA-IGly26Arg elicited macrophage activation, thus stimulating local chronic inflammation. Our results strongly suggest that some natural mutations in apoA-I variants elicit protein tendency to aggregate, but in addition the specific interaction of different variants with macrophages may contribute to cellular stress and toxicity in hereditary amyloidosis. PMID:22952757

  11. Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

    PubMed

    Henderson, Clark M; Bollinger, James G; Becker, Jessica O; Wallace, Jennifer M; Laha, Thomas J; MacCoss, Michael J; Hoofnagle, Andrew N

    2017-07-01

    Proteomic analysis of blood proteins in dried blood spots (DBS) is gaining attention as a possible replacement for measurements in plasma/serum collected by venipuncture. We aimed to develop and provisionally validate a nanoflow LC-PRM-MS method for clinical use. We used Skyline to develop a nanoflow LC-PRM-MS method to quantify glycated hemoglobin-β, apolipoprotein A-I, and apolipoprotein B in DBS. Precision, linearity, interferences, and stability were determined and the method was used to analyze samples from 36 human volunteers. The method was compared with clinically validated measurements in paired blood collected via venipuncture. The method was relatively precise for these proteins (10-11% CV) and linear across the normal concentration ranges of these proteins. Interference from high total serum protein concentration (>8 g/dL) was noted for apolipoprotein A-I and apolipoprotein B. Proteins in DBS were stable for 14 days at temperatures below 25°C and trypsinized samples were stable for 48 h at 7°C. There was moderate correlation with clinical methods (r = 0.783-0.858) and significant bias in individual samples. Although the method had adequate precision and linearity for a biomarker, the accuracy compared with clinically validated assays raises concerns regarding the use of DBS compared with venipuncture for clinical use. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. The Conformation of Lipid-Free Human Apolipoprotein A-I in Solution

    PubMed Central

    Pollard, Ricquita D.; Fulp, Brian; Samuel, Michael P.; Sorci-Thomas, Mary G.; Thomas, Michael J.

    2014-01-01

    Apolipoprotein AI (apoA-I) is the principal acceptor of lipids from ATP-binding cassette transporter A1, a process that yields nascent high density lipoproteins. Analysis of lipidated apoA-I conformation yields a belt or twisted belt in which two strands of apoA-I lie antiparallel to one another. In contrast, biophysical studies have suggested that a part of lipid-free apoA-I was arranged in a 4-helix bundle. To understand how lipid-free apoA-I opens from a bundle to a belt while accepting lipid it was necessary to have a more refined model for the conformation of lipid-free apoA-I. This study reports the conformation of lipid-free human apoA-I using lysine-to-lysine chemical cross-linking in conjunction with disulfide cross-linking achieved using selective cysteine mutations. After proteolysis cross-linked peptides were verified by sequencing using tandem mass spectrometry. The resulting structure is compact with roughly 4 helical regions, amino acids 44 through 186, bundled together. C- and N-terminal ends, amino acids 1-43 and 187-243, respectively, are folded such that they lie close to one another. An unusual feature of the molecule is the high degree of connectivity of lysine40 with 6 other lysines, lysines that are close, e.g., lysine59, to distant lysines, e.g., lysine239, that are at the opposite end of the primary sequence. These results are compared and contrasted with other reported conformations for lipid-free human apoA-I and an NMR study of mouse apoA-I. PMID:24308268

  13. Differential regulation of human apolipoprotein AI and high-density lipoprotein by fenofibrate in hapoAI and hapoAI-CIII-AIV transgenic mice.

    PubMed

    Srivastava, Rai Ajit K; He, Shirley; Newton, Roger S

    2011-02-01

    Fenofibrate, a PPAR-α agonist, lowers triglycerides (TG) and raises high-density lipoproteins (HDL-C) in humans. While fenofibrate is very effective in lowering TG, it does not raise HDL-C in humans to the same extent as seen in human apoAI transgenic (hAI-Tg) mice. We studied the mechanism of this discordance using the following compounds as tools: cholic acid that down-regulates human apoAI, and fenofibrate, that elevates hapoAI and HDL-C in hAI-Tg mice. We hypothesized that additional sequences, including apoCIII and AIV genes on chromosome 11, not present in the hapoAI transgene may be responsible for the dampened effect of fibrates on HDL-C seen in humans. For this, hAI-Tg mice with 11kb DNA segment and hapoAI-CIII-AIV-Tg mice with 33kb DNA segment harboring apoCIII and AIV genes were employed. These mice were treated with fenofibrate and cholic acid. Fenofibrate increased apoAI and HDL-C levels, and HDL size in the apoAI-Tg mice via up-regulation of the hapoAI mRNA and increased activity and mRNA of PLTP, respectively. Consistent with earlier findings, cholic acid showed similar effects of lowering HDL-C, and elevating LDL-C in hAI-Tg mice as well as in the hAI-CIII-AIV-Tg mice. Fenofibrate decreased TG and increased HDL size in hAI-CIII-AIV-Tg mice as well, but surprisingly, did not elevate serum levels of hapoAI or hepatic AI mRNA, suggesting that additional sequences not present in the hapoAI transgene (11kb) may be partly responsible for the dampened effect on HDL-C seen in hAI-CIII-AIV-Tg mice. Since hAI-CIII-AIV-Tg mouse mimics fenofibrate effects seen in humans, this transgenic mouse could serve as a better predictive model for screening HDL-C raising compounds. Copyright © 2010 Elsevier B.V. All rights reserved.

  14. Interaction of human apolipoprotein A-I with model membranes exhibiting lipid domains.

    PubMed

    Arnulphi, Cristina; Sánchez, Susana A; Tricerri, M Alejandra; Gratton, Enrico; Jonas, Ana

    2005-07-01

    Several mechanisms for cell cholesterol efflux have been proposed, including membrane microsolubilization, suggesting that the existence of specific domains could enhance the transfer of lipids to apolipoproteins. In this work isothermal titration calorimetry, circular dichroism spectroscopy, and two-photon microscopy are used to study the interaction of lipid-free apolipoprotein A-I (apoA-I) with small unilamellar vesicles (SUVs) of 1-palmitoyl, 2-oleoyl phosphatidylcholine (POPC) and sphingomyelin (SM), with and without cholesterol. Below 30 degrees C the calorimetric results show that apoA-I interaction with POPC/SM SUVs produces an exothermic reaction, characterized as nonclassical hydrophobic binding. The heat capacity change (DeltaCp degrees ) is small and positive, whereas it was larger and negative for pure POPC bilayers, in the absence of SM. Inclusion of cholesterol in the membranes induces changes in the observed thermodynamic pattern of binding and counteracts the formation of alpha-helices in the protein. Above 30 degrees C the reactions are endothermic. Giant unilamellar vesicles (GUVs) of identical composition to the SUVs, and two-photon fluorescence microscopy techniques, were utilized to further characterize the interaction. Fluorescence imaging of the GUVs indicates coexistence of lipid domains under 30 degrees C. Binding experiments and Laurdan generalized-polarization measurements suggest that there is no preferential binding of the labeled apoA-I to any particular domain. Changes in the content of alpha-helix, binding, and fluidity data are discussed in the framework of the thermodynamic parameters.

  15. Effects of red grape juice consumption on high density lipoprotein-cholesterol, apolipoprotein AI, apolipoprotein B and homocysteine in healthy human volunteers.

    PubMed

    Khadem-Ansari, Mohammad H; Rasmi, Yousef; Ramezani, Fatemeh

    2010-01-01

    It has suggested that grape juice consumption has lipid- lowering effect and it is associated with a decreased risk of heart disease. We aimed to evaluate the effects of red grape juice (RGj) consumption on high density lipoprotein-cholesterol (HDL-C), apolipoprotein AI (apoAI), apolipoprotein B (apoB) and homocysteine (Hcy) levels in healthy human volunteers. Twenty six healthy and nonsmoking males, aged between 25-60 years, who were under no medication asked to consume 150 ml of RGj twice per day for one month. Serum HDL-C, apoAI, apoB and plasma Hcy levels were measured before and after one month RGj consumption. HDL-C levels after RGj consumption were significantly higher than the corresponding levels before the RGj consumption (41.44 ± 4.50 and 44.37 ± 4.30 mg/dl; P<0.0001). Also, apoB was significantly increased after RGj consumption (149.0 ± 22.35 and 157.19 ± 18.60 mg/dl; P<0.002). But apoAI levels were not changed significantly before and after of RGj consumption (154.27 ± 21.55 and 155.35 ± 21.07 mg/dl; P>0.05). Hcy levels were decreased after RGj consumption (7.70 ± 2.80 and 6.20 ± 2.30 µmol/l; P<0.001). The present study demonstrates that RGj consumption can significantly increase serum HDL-C levels and decrease Hcy levels. These findings may have important implications for the prevention of atherosclerosis in healthy individuals.

  16. Interaction of Human Apolipoprotein A-I with Model Membranes Exhibiting Lipid Domains

    PubMed Central

    Arnulphi, Cristina; Sánchez, Susana A.; Tricerri, M. Alejandra; Gratton, Enrico; Jonas, Ana

    2005-01-01

    Several mechanisms for cell cholesterol efflux have been proposed, including membrane microsolubilization, suggesting that the existence of specific domains could enhance the transfer of lipids to apolipoproteins. In this work isothermal titration calorimetry, circular dichroism spectroscopy, and two-photon microscopy are used to study the interaction of lipid-free apolipoprotein A-I (apoA-I) with small unilamellar vesicles (SUVs) of 1-palmitoyl, 2-oleoyl phosphatidylcholine (POPC) and sphingomyelin (SM), with and without cholesterol. Below 30°C the calorimetric results show that apoA-I interaction with POPC/SM SUVs produces an exothermic reaction, characterized as nonclassical hydrophobic binding. The heat capacity change (ΔCp°) is small and positive, whereas it was larger and negative for pure POPC bilayers, in the absence of SM. Inclusion of cholesterol in the membranes induces changes in the observed thermodynamic pattern of binding and counteracts the formation of α-helices in the protein. Above 30°C the reactions are endothermic. Giant unilamellar vesicles (GUVs) of identical composition to the SUVs, and two-photon fluorescence microscopy techniques, were utilized to further characterize the interaction. Fluorescence imaging of the GUVs indicates coexistence of lipid domains under 30°C. Binding experiments and Laurdan generalized-polarization measurements suggest that there is no preferential binding of the labeled apoA-I to any particular domain. Changes in the content of α-helix, binding, and fluidity data are discussed in the framework of the thermodynamic parameters. PMID:15849246

  17. High yield of recombinant human Apolipoprotein A-I expressed in Pichia pastoris by using mixed-mode chromatography.

    PubMed

    Narasimhan Janakiraman, Vignesh; Noubhani, Abdelmajid; Venkataraman, Krishnan; Vijayalakshmi, Mookambeswaran; Santarelli, Xavier

    2016-01-01

    A vast majority of the cardioprotective properties exhibited by High-Density Lipoprotein (HDL) is mediated by its major protein component Apolipoprotein A-I (ApoA1). In order to develop a simplified bioprocess for producing recombinant human Apolipoprotein A-I (rhApoA1) in its near-native form, rhApoA1was expressed without the use of an affinity tag in view of its potential therapeutic applications. Expressed in Pichia pastoris at expression levels of 58.2 mg ApoA1 per litre of culture in a reproducible manner, the target protein was purified by mixed-mode chromatography using Capto™ MMC ligand with a purity and recovery of 84% and 68%, respectively. ApoA1 purification was scaled up to Mixed-mode Expanded Bed Adsorption chromatography to establish an 'on-line' process for the efficient capture of rhApoA1 directly from the P. pastoris expression broth. A polishing step using anion exchange chromatography enabled the recovery of ApoA1 up to 96% purity. Purified ApoA1 was identified and verified by RPLC-ESI-Q-TOF mass spectrometry. This two-step process would reduce processing times and therefore costs in comparison to the twelve-step procedure currently used for recovering rhApoA1 from P. pastoris. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Novel Pathways of Apolipoprotein A-I Metabolism in High-Density Lipoprotein of Different Sizes in Humans.

    PubMed

    Mendivil, Carlos O; Furtado, Jeremy; Morton, Allyson M; Wang, Liyun; Sacks, Frank M

    2016-01-01

    A prevailing concept is that high-density lipoprotein (HDL) is secreted into the systemic circulation as a small mainly discoidal particle, which expands progressively and becomes spherical by uptake and esterification of cellular cholesterol and then contracts by cholesterol ester delivery to the liver, a process known as reverse cholesterol transport, thought to be impaired in people with low HDL cholesterol (HDLc). This metabolic framework has not been established in humans. We studied the metabolism of apolipoprotein A-I in 4 standard HDL sizes by endogenous isotopic labeling in 6 overweight adults with low HDLc and in 6 adults with normal body weight with high plasma HDLc. Contrary to expectation, HDL was secreted into the circulation in its entire size distribution from very small to very large similarly in both groups. Very small (prebeta) HDL comprised only 8% of total apolipoprotein A-I secretion. Each HDL subfraction circulated mostly within its secreted size range for 1 to 4 days and then was cleared. Enlargement of very small and medium to large and very large HDL and generation of very small from medium HDL were minor metabolic pathways. Prebeta HDL was cleared slower, whereas medium, large, and very large HDL were cleared faster in the low HDLc group. A new model is proposed from these results in which HDL is metabolized in plasma mainly within several discrete, stable sizes across the common range of HDLc concentrations. © 2015 American Heart Association, Inc.

  19. Apolipoprotein A-I structural organization in high density lipoproteins isolated from human plasma

    PubMed Central

    Huang, Rong; Gangani D. Silva, R. A.; Jerome, W. Gray; Kontush, Anatol; Chapman, M. John; Curtiss, Linda K.; Hodges, Timothy J.; Davidson, W. Sean

    2010-01-01

    High density lipoproteins (HDL) mediate cholesterol transport and protection from cardiovascular disease. Although synthetic HDLs have been studied for 30 years, the structure of human plasma-derived HDL, and its major protein apolipoprotein (apo)A-I, is unknown. We separated normal human HDL into 5 density subfractions and then further isolated those containing predominantly apoA-I (LpA-I). Using cross-linking chemistry and mass spectrometry, we found that apoA-I adopts a structural framework in these particles that closely mirrors that in synthetic HDL. We adapted established structural models for synthetic HDL to generate the first detailed models of authentic human plasma HDL in which apoA-I adopts a symmetrical cage-like structure. The models suggest that HDL particle size is modulated via a twisting motion of the resident apoA-I molecules. This understanding offers insights into how apoA-I structure modulates HDL function and its interactions with other apolipoproteins. PMID:21399642

  20. Effect of TNF{alpha} on activities of different promoters of human apolipoprotein A-I gene

    SciTech Connect

    Orlov, Sergey V.; Mogilenko, Denis A.; Shavva, Vladimir S.; Dizhe, Ella B.; Ignatovich, Irina A.; Perevozchikov, Andrej P.

    2010-07-23

    Research highlights: {yields} TNF{alpha} stimulates the distal alternative promoter of human apoA-I gene. {yields} TNF{alpha} acts by weakening of promoter competition within apoA-I gene (promoter switching). {yields} MEK1/2 and nuclear receptors PPAR{alpha} and LXRs take part in apoA-I promoter switching. -- Abstract: Human apolipoprotein A-I (ApoA-I) is a major structural and functional protein component of high-density lipoproteins. The expression of the apolipoprotein A-I gene (apoA-I) in hepatocytes is repressed by pro-inflammatory cytokines such as IL-1{beta} and TNF{alpha}. Recently, two novel additional (alternative) promoters for human apoA-I gene have been identified. Nothing is known about the role of alternative promoters in TNF{alpha}-mediated downregulation of apoA-I gene. In this article we report for the first time about the different effects of TNF{alpha} on two alternative promoters of human apoA-I gene. Stimulation of HepG2 cells by TNF{alpha} leads to activation of the distal alternative apoA-I promoter and downregulation of the proximal alternative and the canonical apoA-I promoters. This effect is mediated by weakening of the promoter competition within human apoA-I 5'-regulatory region (apoA-I promoter switching) in the cells treated by TNF{alpha}. The MEK1/2-ERK1/2 cascade and nuclear receptors PPAR{alpha} and LXRs are important for TNF{alpha}-mediated apoA-I promoter switching.

  1. Disruption of human plasma high-density lipoproteins by streptococcal serum opacity factor requires labile apolipoprotein A-I.

    PubMed

    Han, Mikyung; Gillard, Baiba K; Courtney, Harry S; Ward, Kathryn; Rosales, Corina; Khant, Htet; Ludtke, Steven J; Pownall, Henry J

    2009-02-24

    Human plasma high-density lipoproteins (HDL), the primary vehicle for reverse cholesterol transport, are the target of serum opacity factor (SOF), a virulence determinant of Streptococcus pyogenes that turns serum opaque. HDL comprise a core of neutral lipidscholesteryl esters and some triglyceridesurrounded by a surface monolayer of cholesterol, phospholipids, and specialized proteins [apolipoproteins (apos) A-I and A-II]. A HDL is an unstable particle residing in a kinetic trap from which it can escape via chaotropic, detergent, or thermal perturbation. Recombinant (r) SOF catalyzes the transfer of nearly all neutral lipids of approximately 100,000 HDL particles (D approximately 8.5 nm) into a single, large cholesteryl ester-rich microemulsion (CERM; D > 100 nm), leaving a new HDL-like particle [neo HDL (D approximately 5.8 nm)] while releasing lipid-free (LF) apo A-I. CERM formation and apo A-I release have similar kinetics, suggesting parallel or rapid consecutive steps. By using complementary physicochemical methods, we have refined the mechanistic model for HDL opacification. According to size exclusion chromatography, a HDL containing nonlabile apo A-I resists rSOF-mediated opacification. On the basis of kinetic cryo-electron microscopy, rSOF (10 nM) catalyzes the conversion of HDL (4 microM) to neo HDL via a stepwise mechanism in which intermediate-sized particles are seen. Kinetic turbidimetry revealed opacification as a rising exponential reaction with a rate constant k of (4.400 +/- 0.004) x 10(-2) min(-1). Analysis of the kinetic data using transition state theory gave an enthalpy (DeltaH()), entropy (DeltaS(++)), and free energy (DeltaG()) of activation of 73.9 kJ/mol, -66.87 J/K, and 94.6 kJ/mol, respectively. The free energy of activation for opacification is nearly identical to that for the displacement of apo A-I from HDL by guanidine hydrochloride. We conclude that apo A-I lability is required for HDL opacification, LF apo A-I desorption is the

  2. Effect of amino acid distribution of amphipathic helical peptide derived from human apolipoprotein A-I on membrane curvature sensing.

    PubMed

    Tanaka, Masafumi; Takamura, Yuki; Kawakami, Toru; Aimoto, Saburo; Saito, Hiroyuki; Mukai, Takahiro

    2013-03-01

    Amphipathic helix, which senses membrane curvature, is of growing interest. Here we explore the effect of amino acid distribution of amphipathic helical peptide derived from the C-terminal region (residues 220-241) of human apolipoprotein (apo) A-I on membrane curvature sensing. This peptide preferred a curved membrane in a manner similar to full-length apoA-I, although its model peptide did not sense membrane curvature. Substitution of several residues both on the polar and non-polar faces of the amphipathic helix had no significant effect on sensing, suggestive of the elaborate molecular architecture in the C-terminal helical region of apoA-I to exert lipid efflux function. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  3. High-Density Lipoprotein Biogenesis: Defining the Domains Involved in Human Apolipoprotein A-I Lipidation.

    PubMed

    Pollard, Ricquita D; Fulp, Brian; Sorci-Thomas, Mary G; Thomas, Michael J

    2016-09-06

    The first step in removing cholesterol from a cell is the ATP-binding cassette transporter 1 (ABCA1)-driven transfer of cholesterol to lipid-free or lipid-poor apolipoprotein A-I (apoA-I), which yields cholesterol-rich nascent high-density lipoprotein (nHDL) that then matures in plasma to spherical, cholesteryl ester-rich HDL. However, lipid-free apoA-I has a three-dimensional (3D) conformation that is significantly different from that of lipidated apoA-I on nHDL. By comparing the lipid-free apoA-I 3D conformation of apoA-I to that of 9-14 nm diameter nHDL, we formulated the hypothetical helical domain transitions that might drive particle formation. To test the hypothesis, ten apoA-I mutants were prepared that contained two strategically placed cysteines several of which could form intramolecular disulfide bonds and others that could not form these bonds. Mass spectrometry was used to identify amino acid sequence and intramolecular disulfide bond formation. Recombinant HDL (rHDL) formation was assessed with this group of apoA-I mutants. ABCA1-driven nHDL formation was measured in four mutants and wild-type apoA-I. The mutants contained cysteine substitutions in one of three regions: the N-terminus, amino acids 34 and 55 (E34C to S55C), central domain amino acids 104 and 162 (F104C to H162C), and the C-terminus, amino acids 200 and 233 (L200C to L233C). Mutants were studied in the locked form, with an intramolecular disulfide bond present, or unlocked form, with the cysteine thiol blocked by alkylation. Only small amounts of rHDL or nHDL were formed upon locking the central domain. We conclude that both the N- and C-terminal ends assist in the initial steps in lipid acquisition, but that opening of the central domain was essential for particle formation.

  4. Novel pathways of apolipoprotein A-I metabolism in HDL of different sizes in humans

    PubMed Central

    Mendivil, Carlos O.; Furtado, Jeremy; Morton, Allyson M.; Wang, Liyun; Sacks, Frank M.

    2015-01-01

    Objective A prevailing concept is that HDL is secreted into the systemic circulation as a small mainly discoidal particle; which expands progressively and becomes spherical by uptake and esterification of cellular cholesterol; and then contracts by cholesterol ester delivery to the liver, a process known as reverse cholesterol transport, thought to be impaired in people with low HDL cholesterol (HDLc). This metabolic framework has not been established in humans. Approach and results We studied the metabolism of apolipoproteinA-I in four standard HDL sizes by endogenous isotopic labeling in six overweight adults with low HDLc and in six adults with normal body weight with high plasma HDLc. Contrary to expectation, HDL was secreted into the circulation in its entire size distribution from very small to very large, similarly in both groups. Very small (prebeta) HDL comprised only 8% of total apoA-I secretion. Each HDL subfraction circulated mostly within its secreted size range for 1–4 days, and then was cleared. Enlargement of very small and medium to large and very large HDL, and generation of very small from medium HDL were minor metabolic pathways. Prebeta HDL was cleared slower whereas medium, large and very large HDL were cleared faster in the low HDLc group. Conclusions A new model is proposed from these results in which HDL is metabolized in plasma mainly within several discrete, stable sizes, across the common range of HDLc concentrations. PMID:26543096

  5. Definition of human apolipoprotein A-I epitopes recognized by autoantibodies present in patients with cardiovascular diseases.

    PubMed

    Teixeira, Priscila Camillo; Ducret, Axel; Ferber, Philippe; Gaertner, Hubert; Hartley, Oliver; Pagano, Sabrina; Butterfield, Michelle; Langen, Hanno; Vuilleumier, Nicolas; Cutler, Paul

    2014-10-10

    Autoantibodies to apolipoprotein A-I (anti-apoA-I IgG) have been shown to be both markers and mediators of cardiovascular disease, promoting atherogenesis and unstable atherosclerotic plaque. Previous studies have shown that high levels of anti-apoA-I IgGs are independently associated with major adverse cardiovascular events in patients with myocardial infarction. Autoantibody responses to apoA-I can be polyclonal and it is likely that more than one epitope may exist. To identify the specific immunoreactive peptides in apoA-I, we have developed a set of methodologies and procedures to isolate, purify, and identify novel apoA-I endogenous epitopes. First, we generated high purity apoA-I from human plasma, using thiophilic interaction chromatography followed by enzymatic digestion specifically at lysine or arginine residues. Immunoreactivity to the different peptides generated was tested by ELISA using serum obtained from patients with acute myocardial infarction and high titers of autoantibodies to native apoA-I. The immunoreactive peptides were further sequenced by mass spectrometry. Our approach successfully identified two novel immunoreactive peptides, recognized by autoantibodies from patients suffering from myocardial infarction, who contain a high titer of anti-apoA-I IgG. The discovery of these epitopes may open innovative prognostic and therapeutic opportunities potentially suitable to improve current cardiovascular risk stratification.

  6. Amyloidogenic Propensity of a Natural Variant of Human Apolipoprotein A-I: Stability and Interaction with Ligands

    PubMed Central

    Rosú, Silvana A.; Rimoldi, Omar J.; Prieto, Eduardo D.; Curto, Lucrecia M.; Delfino, José M.

    2015-01-01

    A number of naturally occurring mutations of human apolipoprotein A-I (apoA-I) have been associated with hereditary amyloidoses. The molecular mechanisms involved in amyloid-associated pathology remain largely unknown. Here we examined the effects of the Arg173Pro point mutation in apoA-I on the structure, stability, and aggregation propensity, as well as on the ability to bind to putative ligands. Our results indicate that the mutation induces a drastic loss of stability, and a lower efficiency to bind to phospholipid vesicles at physiological pH, which could determine the observed higher tendency to aggregate as pro-amyloidogenic complexes. Incubation under acidic conditions does not seem to induce significant desestabilization or aggregation tendency, neither does it contribute to the binding of the mutant to sodium dodecyl sulfate. While the binding to this detergent is higher for the mutant as compared to wt apoA-I, the interaction of the Arg173Pro variant with heparin depends on pH, being lower at pH 5.0 and higher than wt under physiological pH conditions. We suggest that binding to ligands as heparin or other glycosaminoglycans could be key events tuning the fine details of the interaction of apoA-I variants with the micro-environment, and probably eliciting the toxicity of these variants in hereditary amyloidoses. PMID:25950566

  7. A facile method for isolation of recombinant human apolipoprotein A-I from E. coli.

    PubMed

    Ikon, Nikita; Shearer, Jennifer; Liu, Jianfang; Tran, Jesse J; Feng, ShiBo; Kamei, Ayako; Beckstead, Jennifer A; Kiss, Robert S; Weers, Paul M; Ren, Gang; Ryan, Robert O

    2017-06-01

    Apolipoprotein (apo) A-I is the major protein component of high-density lipoprotein (HDL) and plays key roles in the Reverse Cholesterol Transport pathway. In the past decade, reconstituted HDL (rHDL) has been employed as a therapeutic agent for treatment of atherosclerosis. The ability of rHDL to promote cholesterol efflux from peripheral cells has been documented to reduce the size of atherosclerotic plaque lesions. However, development of apoA-I rHDL-based therapeutics for human use requires a cost effective process to generate an apoA-I product that meets "Good Manufacturing Practice" standards. Methods available for production and isolation of unmodified recombinant human apoA-I at scale are cumbersome, laborious and complex. To overcome this obstacle, a streamlined two-step procedure has been devised for isolation of recombinant untagged human apoA-I from E. coli that takes advantage of its ability to re-fold to a native conformation following denaturation. Heat treatment of a sonicated E. coli supernatant fraction induced precipitation of a large proportion of host cell proteins (HCP), yielding apoA-I as the major soluble protein. Reversed-phase HPLC of this material permitted recovery of apoA-I largely free of HCP and endotoxin. Purified apoA-I possessed α-helix secondary structure, formed rHDL upon incubation with phospholipid and efficiently promoted cholesterol efflux from cholesterol loaded J774 macrophages. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. A facile method for isolation of recombinant human apolipoprotein A-I from E. coli

    DOE PAGES

    Ikon, Nikita; Shearer, Jennifer; Liu, Jianfang; ...

    2017-03-20

    Apolipoprotein (apo) A-I is the major protein component of high-density lipoprotein (HDL) and plays key roles in the Reverse Cholesterol Transport pathway. In the past decade, reconstituted HDL (rHDL) has been employed as a therapeutic agent for treatment of atherosclerosis. The ability of rHDL to promote cholesterol efflux from peripheral cells has been documented to reduce the size of atherosclerotic plaque lesions. However, development of apoA-I rHDL-based therapeutics for human use requires a cost effective process to generate an apoA-I product that meets “Good Manufacturing Practice” standards. Methods available for production and isolation of unmodified recombinant human apoA-I at scalemore » are cumbersome, laborious and complex. To overcome this obstacle, a streamlined two-step procedure has been devised for isolation of recombinant untagged human apoA-I from E. coli that takes advantage of its ability to re-fold to a native conformation following denaturation. Heat treatment of a sonicated E. coli supernatant fraction induced precipitation of a large proportion of host cell proteins (HCP), yielding apoA-I as the major soluble protein. Reversed-phase HPLC of this material permitted recovery of apoA-I largely free of HCP and endotoxin. In conclusion, purified apoA-I possessed α-helix secondary structure, formed rHDL upon incubation with phospholipid and efficiently promoted cholesterol efflux from cholesterol loaded J774 macrophages.« less

  9. Apolipoprotein A-I enhances proliferation of human endothelial progenitor cells and promotes angiogenesis through the cell surface ATP synthase.

    PubMed

    González-Pecchi, Valentina; Valdés, Sara; Pons, Véronique; Honorato, Paula; Martinez, Laurent O; Lamperti, Liliana; Aguayo, Claudio; Radojkovic, Claudia

    2015-03-01

    Human endothelial progenitor cells (hEPC) correspond to a subtype of stem cells which, in the presence of angiogenic stimuli, can be mobilized from bone marrow to circulation and then recruited to the damaged endothelium, where they differentiate into mature endothelial cells. High-density lipoproteins (HDL) increase the level and functionality (proliferation, migration, differentiation, angiogenesis capacity) of circulating hEPC; however, the contribution of receptors for HDL and/or apolipoprotein A-I (apoA-I), the main HDL apolipoprotein, in these effects is still unclear. On mature endothelial cells, the cell surface F1-ATP synthase has been previously characterized as a high affinity receptor of apoA-I, whereas the scavenger receptor SR-BI mainly binds with fully lipidated HDL and displays a poor affinity for lipid-free apoA-I. Furthermore, it was shown that apoA-I binding to surface ATP synthase on mature endothelial cells promotes cell proliferation, whereas inhibits apoptosis. In this work, we aimed to determine the effect of apoA-I in the proliferation and the angiogenic capacity of early hEPC, and the contribution of the cell surface ATP synthase in these events. We first evidenced that early hEPC express the ATP synthase at the surface of nonpermeabilized cells, where it is not colocalized with MitoTracker, a mitochondria marker. ApoA-I (50 μg/mL) increases hEPC proliferation (+14.5%, p<0.001) and potentiates the effect of hEPC on a cellular model of angiogenesis, with an increase of +31% (p<0.01) in branch point counting and in tubule length. These effects of apoA-I were totally reversed in the presence of ATP synthase inhibitors, such as IF1 or oligomycin, whereas the inhibition of the HDL receptor, SR-BI, partially inhibits these events. These results provide the first evidence that surface ATP synthase is expressed on early hEPC, where it mediates apoA-I effects in hEPC proliferation and in angiogenesis. This knowledge could be helpful for future

  10. Comparative models for human apolipoprotein A-I bound to lipid in discoidal high-density lipoprotein particles.

    PubMed

    Klon, Anthony E; Segrest, Jere P; Harvey, Stephen C

    2002-09-10

    We have constructed a series of models for apolipoprotein A-I (apo A-I) bound to discoidal high-density lipoprotein (HDL) particles, based upon the molecular belt model [Segrest, J. P., et al. (1999) J. Biol. Chem. 274, 31755-31758] and helical hairpin models [Rogers, D. P., et al. (1998) Biochemistry 37, 11714-11725], and compared these with picket fence models [Phillips, J. C., et al. (1997) Biophys. J. 73, 2337-2346]. Molecular belt models for discoidal HDL particles with differing diameters are presented, illustrating that the belt model can explain the discrete changes in HDL particle size observed experimentally. Hairpin models are discussed for the binding of apo A-I to discoidal HDL particles with diameters identical to those for the molecular belt model. Two models are presented for the binding of three monomers of apo A-I to a 150 A diameter discoidal HDL particle. In one model, two monomers of apo A-I bind to the exterior of the HDL particle in an antiparallel belt, with a third monomer of apo A-I bound to the disk in a hairpin conformation. In the second model, all three monomers of apo A-I are bound to the discoidal HDL particle in a hairpin conformation. Previously published experimental data for each model are reviewed, with FRET favoring either the belt or hairpin models over the picket fence models for HDL particles with diameters of 105 A. Naturally occurring mutations appear to favor the belt model for the 105 A particles, while the 150 A HDL particles favor the presence of at least one hairpin.

  11. Carboxyl-Terminal Cleavage of Apolipoprotein A-I by Human Mast Cell Chymase Impairs Its Anti-Inflammatory Properties.

    PubMed

    Nguyen, Su Duy; Maaninka, Katariina; Lappalainen, Jani; Nurmi, Katariina; Metso, Jari; Öörni, Katariina; Navab, Mohamad; Fogelman, Alan M; Jauhiainen, Matti; Lee-Rueckert, Miriam; Kovanen, Petri T

    2016-02-01

    Apolipoprotein A-I (apoA-I) has been shown to possess several atheroprotective functions, including inhibition of inflammation. Protease-secreting activated mast cells reside in human atherosclerotic lesions. Here we investigated the effects of the neutral proteases released by activated mast cells on the anti-inflammatory properties of apoA-I. Activation of human mast cells triggered the release of granule-associated proteases chymase, tryptase, cathepsin G, carboxypeptidase A, and granzyme B. Among them, chymase cleaved apoA-I with the greatest efficiency and generated C-terminally truncated apoA-I, which failed to bind with high affinity to human coronary artery endothelial cells. In tumor necrosis factor-α-activated human coronary artery endothelial cells, the chymase-cleaved apoA-I was unable to suppress nuclear factor-κB-dependent upregulation of vascular cell adhesion molecule-1 (VCAM-1) and to block THP-1 cells from adhering to and transmigrating across the human coronary artery endothelial cells. Chymase-cleaved apoA-I also had an impaired ability to downregulate the expression of tumor necrosis factor-α, interleukin-1β, interleukin-6, and interleukin-8 in lipopolysaccharide-activated GM-CSF (granulocyte-macrophage colony-stimulating factor)- and M-CSF (macrophage colony-stimulating factor)-differentiated human macrophage foam cells and to inhibit reactive oxygen species formation in PMA (phorbol 12-myristate 13-acetate)-activated human neutrophils. Importantly, chymase-cleaved apoA-I showed reduced ability to inhibit lipopolysaccharide-induced inflammation in vivo in mice. Treatment with chymase blocked the ability of the apoA-I mimetic peptide L-4F, but not of the protease-resistant D-4F, to inhibit proinflammatory gene expression in activated human coronary artery endothelial cells and macrophage foam cells and to prevent reactive oxygen species formation in activated neutrophils. The findings identify C-terminal cleavage of apoA-I by human mast

  12. Carboxyl-Terminal Cleavage of Apolipoprotein A-I by Human Mast Cell Chymase Impairs Its Anti-Inflammatory Properties

    PubMed Central

    Nguyen, Su Duy; Maaninka, Katariina; Lappalainen, Jani; Nurmi, Katariina; Metso, Jari; Öörni, Katariina; Navab, Mohamad; Fogelman, Alan M.; Jauhiainen, Matti; Lee-Rueckert, Miriam

    2016-01-01

    Objective— Apolipoprotein A-I (apoA-I) has been shown to possess several atheroprotective functions, including inhibition of inflammation. Protease-secreting activated mast cells reside in human atherosclerotic lesions. Here we investigated the effects of the neutral proteases released by activated mast cells on the anti-inflammatory properties of apoA-I. Approach and Results— Activation of human mast cells triggered the release of granule-associated proteases chymase, tryptase, cathepsin G, carboxypeptidase A, and granzyme B. Among them, chymase cleaved apoA-I with the greatest efficiency and generated C-terminally truncated apoA-I, which failed to bind with high affinity to human coronary artery endothelial cells. In tumor necrosis factor-α–activated human coronary artery endothelial cells, the chymase-cleaved apoA-I was unable to suppress nuclear factor-κB–dependent upregulation of vascular cell adhesion molecule-1 (VCAM-1) and to block THP-1 cells from adhering to and transmigrating across the human coronary artery endothelial cells. Chymase-cleaved apoA-I also had an impaired ability to downregulate the expression of tumor necrosis factor-α, interleukin-1β, interleukin-6, and interleukin-8 in lipopolysaccharide-activated GM-CSF (granulocyte-macrophage colony-stimulating factor)– and M-CSF (macrophage colony-stimulating factor)–differentiated human macrophage foam cells and to inhibit reactive oxygen species formation in PMA (phorbol 12-myristate 13-acetate)–activated human neutrophils. Importantly, chymase-cleaved apoA-I showed reduced ability to inhibit lipopolysaccharide-induced inflammation in vivo in mice. Treatment with chymase blocked the ability of the apoA-I mimetic peptide L-4F, but not of the protease-resistant D-4F, to inhibit proinflammatory gene expression in activated human coronary artery endothelial cells and macrophage foam cells and to prevent reactive oxygen species formation in activated neutrophils. Conclusions— The

  13. D-4F, an apolipoprotein A-I mimetic, inhibits TGF-β1 induced epithelial-mesenchymal transition in human alveolar epithelial cell.

    PubMed

    You, Jia; Wang, Jintao; Xie, Linshen; Zhu, Chengwen; Xiong, Jingyuan

    2016-10-01

    Emerging evidences support that transforming growth factor β1 (TGF-β1) induced epithelial-mesenchymal transition (EMT) participates in the pathogenesis of pulmonary fibrosis and asthmatic airway remodeling. Recent studies demonstrated that apolipoprotein A-I (Apo A-I) is the only known substance that can resolve established pulmonary fibrotic nodules, and Apo A-I mimetic D-4F (a synthetic polypeptide consisting of 18 amino acids) plays an inhibitory role in murine asthmatic model. However, cellular mechanisms for such therapeutic effects of Apo A-I and D-4F remain to be elucidated. This study evaluated the effects of D-4F on TGF-β1 induced EMT in human type II alveolar epithelial cell line A549. A549 cells treated with 10ng/ml of TGF-β1 manifested distinct EMT, including fibroblastic morphological changes, down-regulation of epithelial marker E-cadherin and up-regulation of mesenchymal marker vimentin. These EMT related changes were all inhibited by D-4F in a concentration dependent manner. Transcriptional investigation demonstrated clearly that D-4F dose-dependently compensated for the reduced E-cadherin mRNA level and the increased vimentin mRNA level in TGF-β1 treated A549 cells. Translational analysis revealed that D-4F significantly reversed the TGF-β1 induced changes of E-cadherin and vimentin levels. These results suggested that D-4F inhibits TGF-β1 induced EMT in human alveolar epithelial cell. Given the functional similarities between D-4F and Apo A-I, it is speculated that D-4F and Apo A-I are able to exert possible anti-fibrotic and anti-asthmatic effects via inhibiting alveolar EMT, and D-4F may possess beneficial clinical potential for patients suffering from pulmonary fibrosis and asthma. Copyright © 2016 Elsevier GmbH. All rights reserved.

  14. Featured Article: Alterations of lecithin cholesterol acyltransferase activity and apolipoprotein A-I functionality in human sickle blood

    PubMed Central

    Borja, Mark S; Borda, Mauricio; Larkin, Sandra K; Kuypers, Frans A

    2016-01-01

    In sickle cell disease (SCD) cholesterol metabolism appears dysfunctional as evidenced by abnormal plasma cholesterol content in a subpopulation of SCD patients. Specific activity of the high density lipoprotein (HDL)-bound lecithin cholesterol acyltransferase (LCAT) enzyme, which catalyzes esterification of cholesterol, and generates lysoPC (LPC) was significantly lower in sickle plasma compared to normal. Inhibitory amounts of LPC were present in sickle plasma, and the red blood cell (RBC) lysophosphatidylcholine acyltransferase (LPCAT), essential for the removal of LPC, displayed a broad range of activity. The functionality of sickle HDL appeared to be altered as evidenced by a decreased HDL–Apolipoprotein A-I exchange in sickle plasma as compared to control. Increased levels of oxidized proteins including ApoA-I were detected in sickle plasma. In vitro incubation of sickle plasma with washed erythrocytes affected the ApoA-I-exchange supporting the view that the RBC blood compartment can affect cholesterol metabolism in plasma. HDL functionality appeared to decrease during acute vaso-occlusive episodes in sickle patients and was associated with an increase of secretory PLA2, a marker for increased inflammation. Simvastatin treatment to improve the anti-inflammatory function of HDL did not ameliorate HDL–ApoA-I exchange in sickle patients. Thus, the cumulative effect of an inflammatory and highly oxidative environment in sickle blood contributes to a decrease in cholesterol esterification and HDL function, related to hypocholesterolemia in SCD. PMID:27354333

  15. Featured Article: Alterations of lecithin cholesterol acyltransferase activity and apolipoprotein A-I functionality in human sickle blood.

    PubMed

    Soupene, Eric; Borja, Mark S; Borda, Mauricio; Larkin, Sandra K; Kuypers, Frans A

    2016-11-01

    In sickle cell disease (SCD) cholesterol metabolism appears dysfunctional as evidenced by abnormal plasma cholesterol content in a subpopulation of SCD patients. Specific activity of the high density lipoprotein (HDL)-bound lecithin cholesterol acyltransferase (LCAT) enzyme, which catalyzes esterification of cholesterol, and generates lysoPC (LPC) was significantly lower in sickle plasma compared to normal. Inhibitory amounts of LPC were present in sickle plasma, and the red blood cell (RBC) lysophosphatidylcholine acyltransferase (LPCAT), essential for the removal of LPC, displayed a broad range of activity. The functionality of sickle HDL appeared to be altered as evidenced by a decreased HDL-Apolipoprotein A-I exchange in sickle plasma as compared to control. Increased levels of oxidized proteins including ApoA-I were detected in sickle plasma. In vitro incubation of sickle plasma with washed erythrocytes affected the ApoA-I-exchange supporting the view that the RBC blood compartment can affect cholesterol metabolism in plasma. HDL functionality appeared to decrease during acute vaso-occlusive episodes in sickle patients and was associated with an increase of secretory PLA2, a marker for increased inflammation. Simvastatin treatment to improve the anti-inflammatory function of HDL did not ameliorate HDL-ApoA-I exchange in sickle patients. Thus, the cumulative effect of an inflammatory and highly oxidative environment in sickle blood contributes to a decrease in cholesterol esterification and HDL function, related to hypocholesterolemia in SCD. © 2016 by the Society for Experimental Biology and Medicine.

  16. Human macrophage cathepsin B-mediated C-terminal cleavage of apolipoprotein A-I at Ser228 severely impairs antiatherogenic capacity.

    PubMed

    Dinnes, Donna Lee M; White, Melanie Y; Kockx, Maaike; Traini, Mathew; Hsieh, Victar; Kim, Mi-Jurng; Hou, Liming; Jessup, Wendy; Rye, Kerry-Anne; Thaysen-Andersen, Morten; Cordwell, Stuart J; Kritharides, Leonard

    2016-12-01

    Apolipoprotein A-I (apoA-I) is the major component of HDL and central to the ability of HDL to stimulate ATP-binding cassette transporter A1 (ABCA1)-dependent, antiatherogenic export of cholesterol from macrophage foam cells, a key player in the pathology of atherosclerosis. Cell-mediated modifications of apoA-I, such as chlorination, nitration, oxidation, and proteolysis, can impair its antiatherogenic function, although it is unknown whether macrophages themselves contribute to such modifications. To investigate this, human monocyte-derived macrophages (HMDMs) were incubated with human apoA-I under conditions used to induce cholesterol export. Two-dimensional gel electrophoresis and Western blot analysis identified that apoA-I is cleaved (∼20-80%) by HMDMs in a time-dependent manner, generating apoA-I of lower MW and isoelectric point. Mass spectrometry analysis identified a novel C-terminal cleavage site of apoA-I between Ser(228)-Phe(229) Recombinant apoA-I truncated at Ser(228) demonstrated profound loss of capacity to solubilize lipid and to promote ABCA1-dependent cholesterol efflux. Protease inhibitors, small interfering RNA knockdown in HMDMs, mass spectrometry analysis, and cathepsin B activity assays identified secreted cathepsin B as responsible for apoA-I cleavage at Ser(228) Importantly, C-terminal cleavage of apoA-I was also detected in human carotid plaque. Cleavage at Ser(228) is a novel, functionally important post-translational modification of apoA-I mediated by HMDMs that limits the antiatherogenic properties of apoA-I.-Dinnes, D. L. M., White, M. Y., Kockx, M., Traini, M., Hsieh, V., Kim, M.-J., Hou, L., Jessup, W., Rye, K.-A., Thaysen-Andersen, M., Cordwell, S. J., Kritharides, L. Human macrophage cathepsin B-mediated C-terminal cleavage of apolipoprotein A-I at Ser(228) severely impairs antiatherogenic capacity. © FASEB.

  17. Serum opacity factor unmasks human plasma high-density lipoprotein instability via selective delipidation and apolipoprotein A-I desorption.

    PubMed

    Gillard, Baiba K; Courtney, Harry S; Massey, John B; Pownall, Henry J

    2007-11-13

    Human plasma high-density lipoproteins (HDL) are important vehicles in reverse cholesterol transport, the cardioprotective mechanism by which peripheral tissue-cholesterol is transported to the liver for disposal. HDL is the target of serum opacity factor (SOF), a substance produced by Streptococcus pyogenes that turns mammalian serum cloudy. Using a recombinant (r) SOF, we studied opacification and its mechanism. rSOF catalyzes the partial disproportionation of HDL into a cholesteryl ester-rich microemulsion (CERM) and a new HDL-like particle, neo HDL, with the concomitant release of lipid-free (LF)-apo A-I. Opacification is unique; rSOF transfers apo E and nearly all neutral lipids of approximately 100,000 HDL particles into a single large CERM whose size increases with HDL-CE content (r approximately 100-250 nm) leaving a neo HDL that is enriched in PL (41%) and protein (48%), especially apo A-II. rSOF is potent; within 30 min at 37 degrees C, 10 nM rSOF opacifies 4 microM HDL. At respective low and high physiological HDL concentrations, LF-apo A-I is monomeric and tetrameric. CERM formation and apo A-I release have similar kinetics suggesting parallel or rapid sequential steps. According to the reaction products and kinetics, rSOF is a heterodivalent fusogenic protein that uses a docking site to displace apo A-I and bind to exposed CE surfaces on HDL; the resulting rSOF-HDL complex recruits additional HDL with its binding-delipidation site and through multiple fusion steps forms a CERM. rSOF may be a clinically useful and novel modality for improving reverse cholesterol transport. With apo E and a high CE content, CERM could transfer large amounts of cholesterol to the liver for disposal via the LDL receptor; neo HDL is likely a better acceptor of cellular cholesterol than HDL; LF-apo A-I could enhance efflux via the ATP-binding casette transporter ABCA1.

  18. Apolipoprotein A-I: A Molecule of Diverse Function.

    PubMed

    Mangaraj, Manaswini; Nanda, Rachita; Panda, Suchismita

    2016-07-01

    Apolipoprotein A-I (apo A-I) an indispensable component and a major structural protein of high-density lipoprotein (HDL), plays a vital role in reverse cholesterol transport and cellular cholesterol homeostasis since its identification. Its multifunctional role in immunity, inflammation, apoptosis, viral, bacterial infection etc. has crossed its boundary of its potential of protecting cardiovascular system and lowering cardiovascular disease risk, attributing HDL to be known as a protective fat removal particle. Its structural homology with prostacyclin stabilization factor has contributed to its anti-clotting and anti-aggregatory effect on platelet which has potentiated its cardio-protective role as well as its therapeutic efficacy against Alzheimer's disease. The binding affinity and neutralising action against endotoxin lipopolysaccharide, reduces the toxic manifestations of septic shock. As a negative acute phase protein, it blocks T-cell signalling of macrophages. However the recently identified anti-tumor activity of apo A-I has been highlighted in various models of melanoma, lung cancer, ovarian cancer, lymphoblastic leukaemia, gastric as well as pancreatic cancers. These cancer fighting effects are directed towards regression of tumor size and distant metastasis by its immuno modulatory activity as well as its clearing effect on serum lysophospholipids. This lowering effect on lysophospholipid concentration is utilized by apo A-I mimetic peptides to be used in retarding tumor cell proliferation and as a potential cancer therapeutic agent. Not only that, it inhibits the tumor associated neo-angiogenesis as well as brings down the matrix degrading enzymes associated with tumor metastasis. However this efficient therapeutic potential of apo A-I as an anti tumor agent awaits further future experimental studies in humans.

  19. Intravenously injected human apolipoprotein A-I rapidly enters the central nervous system via the choroid plexus.

    PubMed

    Stukas, Sophie; Robert, Jerome; Lee, Michael; Kulic, Iva; Carr, Michael; Tourigny, Katherine; Fan, Jianjia; Namjoshi, Dhananjay; Lemke, Kalistyne; DeValle, Nicole; Chan, Jeniffer; Wilson, Tammy; Wilkinson, Anna; Chapanian, Rafi; Kizhakkedathu, Jayachandran N; Cirrito, John R; Oda, Michael N; Wellington, Cheryl L

    2014-11-12

    Brain lipoprotein metabolism is dependent on lipoprotein particles that resemble plasma high-density lipoproteins but that contain apolipoprotein (apo) E rather than apoA-I as their primary protein component. Astrocytes and microglia secrete apoE but not apoA-I; however, apoA-I is detectable in both cerebrospinal fluid and brain tissue lysates. The route by which plasma apoA-I enters the central nervous system is unknown. Steady-state levels of murine apoA-I in cerebrospinal fluid and interstitial fluid are 0.664 and 0.120 μg/mL, respectively, whereas brain tissue apoA-I is ≈10% to 15% of its levels in liver. Recombinant, fluorescently tagged human apoA-I injected intravenously into mice localizes to the choroid plexus within 30 minutes and accumulates in a saturable, dose-dependent manner in the brain. Recombinant, fluorescently tagged human apoA-I accumulates in the brain for 2 hours, after which it is eliminated with a half-life of 10.3 hours. In vitro, human apoA-I is specifically bound, internalized, and transported across confluent monolayers of primary human choroid plexus epithelial cells and brain microvascular endothelial cells. Following intravenous injection, recombinant human apoA-I rapidly localizes predominantly to the choroid plexus. Because apoA-I mRNA is undetectable in murine brain, our results suggest that plasma apoA-I, which is secreted from the liver and intestine, gains access to the central nervous system primarily by crossing the blood-cerebrospinal fluid barrier via specific cellular mediated transport, although transport across the blood-brain barrier may also contribute to a lesser extent. © 2014 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

  20. Docosahexaenoic acid suppresses apolipoprotein A-I gene expression through hepatocyte nuclear factor-3beta

    USDA-ARS?s Scientific Manuscript database

    BACKGROUND: Dietary fish-oil supplementation has been shown in human kinetic studies to lower the production rate of apolipoprotein (apo) A-I, the major protein component of HDL. The underlying mechanism responsible for this effect is not fully understood. OBJECTIVE: We investigated the effect and...

  1. Reagent or myeloperoxidase-generated hypochlorite affects discrete regions in lipid-free and lipid-associated human apolipoprotein A-I.

    PubMed Central

    Bergt, C; Oettl, K; Keller, W; Andreae, F; Leis, H J; Malle, E; Sattler, W

    2000-01-01

    We have previously shown that the modification of high-density lipoprotein subclass 3 (HDL(3)) by HOCl transformed an anti-atherogenic lipoprotein into a high-uptake form for macrophages and caused a significant impairment of cholesterol efflux capacity [Panzenboeck, Raitmayer, Reicher, Lindner, Glatter, Malle and Sattler (1997) J. Biol. Chem. 272, 29711-29720]. To elucidate the consequences of treatment with OCl(-) on distinct regions in apolipoprotein A-I (apo A-I), lipid-free and lipid-associated apo A-I were modified with increasing molar ratios of NaOCl or HOCl generated by the myeloperoxidase/H(2)O(2)/Cl(-) system. CD analysis revealed a pronounced decrease in alpha-helicity for lipid-free apo A-I modified by NaOCl, whereas lipid-associated apo A-I was less affected. The modification of apo A-I by NaOCl (molar oxidant-to-lipoprotein ratio 6:1) resulted in the formation of two distinct oxidized forms of apo A-I with molecular masses 32 or 48 atomic mass units (a.m.u.) higher than that of native apo A-I, indicating the addition of two or three oxygen atoms to the native protein. HPLC analysis of tryptic digests obtained from lipid-free and lipid-associated apo A-I modified with increasing oxidant-to-apolipoprotein molar ratios revealed a concentration-dependent modification of apo A-I: at a low molar oxidant-to-lipoprotein ratio (5:1) the peaks corresponding to the methionine-containing tryptic peptides T11 (residues 84-88), T16 (residues 108-116) and T22 (residues 141-149), located in the central region of apo A-I, disappeared. Their loss was accompanied by the formation of three oxidation products with a molecular mass 16 a.m.u. higher than that of the native peptides. This indicates the addition of oxygen, most probably caused by the oxidation of Met(86), Met(112) and Met(148) to the corresponding methionine sulphoxides. At a molar NaOCl-to-apo A-I ratio of 10:1 the disappearance of peptides T1 (residues 1-10), T7 (residues 46-59) and T9 (residues 62-77) was

  2. Human apolipoprotein A-I exerts a prophylactic effect on high-fat diet-induced atherosclerosis via inflammation inhibition in a rabbit model.

    PubMed

    Li, Jiyang; Wang, Weina; Han, Lei; Feng, Meiqing; Lu, Hui; Yang, Li; Hu, Xiangxiang; Shi, Si; Jiang, Shanshan; Wang, Qian; Ye, Li

    2017-02-06

    Apolipoprotein A-I (apoA-I) is the major functional protein fraction of high-density lipoprotein. The prophylactic effect and mechanism of human apoA-I on atherosclerosis (AS) were investigated in a high-fat diet-induced AS rabbit model. The rabbits were injected with apoA-I once a week while fed high-fat diet for 20 weeks. Our results showed that apoA-I could raise the serum level of high-density lipoprotein-cholesterol and reduce those of lipid total cholesterol, triglyceride, and low-density lipoprotein-cholesterol in AS rabbits. Decreased aortic plaque area and aortic injury degree were also observed by Oil Red O staining and HE staining in apoA-I-treated high-fat diet-induced AS rabbits. Further study elucidated that apoA-I could down-regulate the expression of some inflammatory mediators including intercellular adhesion molecule type 1, vascular adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1, tumor necrosis factor-α, interleukin-6 (IL-6), and C-reactive protein in serum and aorta of AS rabbits. In addition, real-time quantitative RT-PCR analyses showed that the apoA-I infusions decreased the mRNA levels of two pro-inflammatory molecules, i.e. nuclear factor kappa B (NF-κB) and cyclooxygenase-2 (COX-2), in aorta of AS rabbits, which was associated with a concomitant reduction in endothelial VCAM-1 and IL-6 mRNA transcription. Together, our results support the atheroprotective and prophylactic role of apoA-I in vivo, and this activity may be correlated with its anti-inflammatory effect. © The Author 2017. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Cumulative Brain Injury from Motor Vehicle-Induced Whole-Body Vibration and Prevention by Human Apolipoprotein A-I Molecule Mimetic (4F) Peptide (an Apo A-I Mimetic).

    PubMed

    Yan, Ji-Geng; Zhang, Lin-ling; Agresti, Michael; Yan, Yuhui; LoGiudice, John; Sanger, James R; Matloub, Hani S; Pritchard, Kirkwood A; Jaradeh, Safwan S; Havlik, Robert

    2015-12-01

    Insidious cumulative brain injury from motor vehicle-induced whole-body vibration (MV-WBV) has not yet been studied. The objective of the present study is to validate whether whole-body vibration for long periods causes cumulative brain injury and impairment of the cerebral function. We also explored a preventive method for MV-WBV injury. A study simulating whole-body vibration was conducted in 72 male Sprague-Dawley rats divided into 9 groups (N = 8): (1) 2-week normal control; (2) 2-week sham control (in the tube without vibration); (3) 2-week vibration (exposed to whole-body vibration at 30 Hz and .5 G acceleration for 4 hours/day, 5 days/week for 2 weeks; vibration parameters in the present study are similar to the most common driving conditions); (4) 4-week sham control; (5) 4-week vibration; (6) 4-week vibration with human apolipoprotein A-I molecule mimetic (4F)-preconditioning; (7) 8-week sham control; (8) 8-week vibration; and (9) 8-week 4F-preconditioning group. All the rats were evaluated by behavioral, physiological, and histological studies of the brain. Brain injury from vibration is a cumulative process starting with cerebral vasoconstriction, squeezing of the endothelial cells, increased free radicals, decreased nitric oxide, insufficient blood supply to the brain, and repeated reperfusion injury to brain neurons. In the 8-week vibration group, which indicated chronic brain edema, shrunken neuron numbers increased and whole neurons atrophied, which strongly correlated with neural functional impairment. There was no prominent brain neuronal injury in the 4F groups. The present study demonstrated cumulative brain injury from MV-WBV and validated the preventive effects of 4F preconditioning. Copyright © 2015 National Stroke Association. All rights reserved.

  4. Conformation and Lipid Binding of a C-Terminal (198-243) Peptide of Human Apolipoprotein A-I (apoA-I)†

    PubMed Central

    Zhu, Hongli L.; Atkinson, David

    2008-01-01

    Human apolipoprotein A-I (apoA-I) is the principle apolipoprotein of high-density lipoproteins that are critically involved in reverse cholesterol transport. The intrinsically flexibility of apoA-I has hindered studies of the structural and functional details of the protein. Our strategy is to study peptide models representing different regions of apoA-I. Our previous report on [1-44]apoA-I demonstrated that this N-terminal region is unstructured and folds into ~ 60% α-helix with a moderate lipid binding affinity. We now present details of the conformation and lipid interaction of a C-terminal 46 residue peptide, [198-243]apoA-I, encompassing putative helix repeats 10, 9 and the second half of repeat 8 from the C-terminus of apoA-I. Far ultraviolet circular dichroism spectra show that [198-243] apoA-I is also unfolded in aqueous solution. However, self-association induces ~ 50% α-helix in the peptide. The self-associated peptide exists mainly as a tetramer, as determined by native electrophoresis, cross-linking with glutaraldehyde and unfolding data from circular dichroism (CD) and differential scanning calorimetry (DSC). In the presence of a number of lipid mimicking detergents, above their CMC, ~ 60% α-helix was induced in the peptide. In contrast, SDS, an anionic lipid mimicking detergent, induced helical folding in the peptide at a concentration of ~ 0.003% (~ 100 μM), ~ 70 fold below its typical CMC (0.17–0.23% or 6–8 mM). Both monomeric and tetrameric peptide can solublize dimyristoyl phosphatidyl choline (DMPC) liposomes and fold into ~ 60% α-helix. Fractionation by density gradient ultracentrifugation and visualization by negative staining electromicroscopy, demonstrated that the peptide binds to DMPC with high affinity to form at least two sizes of relatively homogenous discoidal HDL-like particles depending on the initial lipid:peptide ratio. The characteristics (lipid:peptide w/w, diameter and density) of both complexes are similar to those of

  5. Site-specific Nitration of Apolipoprotein A-I at Tyrosine 166 Is Both Abundant within Human Atherosclerotic Plaque and Dysfunctional*

    PubMed Central

    DiDonato, Joseph A.; Aulak, Kulwant; Huang, Ying; Wagner, Matthew; Gerstenecker, Gary; Topbas, Celalettin; Gogonea, Valentin; DiDonato, Anthony J.; Tang, W. H. Wilson; Mehl, Ryan A.; Fox, Paul L.; Plow, Edward F.; Smith, Jonathan D.; Fisher, Edward A.; Hazen, Stanley L.

    2014-01-01

    We reported previously that apolipoprotein A-I (apoA-I) is oxidatively modified in the artery wall at tyrosine 166 (Tyr166), serving as a preferred site for post-translational modification through nitration. Recent studies, however, question the extent and functional importance of apoA-I Tyr166 nitration based upon studies of HDL-like particles recovered from atherosclerotic lesions. We developed a monoclonal antibody (mAb 4G11.2) that recognizes, in both free and HDL-bound forms, apoA-I harboring a 3-nitrotyrosine at position 166 apoA-I (NO2-Tyr166-apoA-I) to investigate the presence, distribution, and function of this modified apoA-I form in atherosclerotic and normal artery wall. We also developed recombinant apoA-I with site-specific 3-nitrotyrosine incorporation only at position 166 using an evolved orthogonal nitro-Tyr-aminoacyl-tRNA synthetase/tRNACUA pair for functional studies. Studies with mAb 4G11.2 showed that NO2-Tyr166-apoA-I was easily detected in atherosclerotic human coronary arteries and accounted for ∼8% of total apoA-I within the artery wall but was nearly undetectable (>100-fold less) in normal coronary arteries. Buoyant density ultracentrifugation analyses showed that NO2-Tyr166-apoA-I existed as a lipid-poor lipoprotein with <3% recovered within the HDL-like fraction (d = 1.063–1.21). NO2-Tyr166-apoA-I in plasma showed a similar distribution. Recovery of NO2-Tyr166-apoA-I using immobilized mAb 4G11.2 showed an apoA-I form with 88.1 ± 8.5% reduction in lecithin-cholesterol acyltransferase activity, a finding corroborated using a recombinant apoA-I specifically designed to include the unnatural amino acid exclusively at position 166. Thus, site-specific nitration of apoA-I at Tyr166 is an abundant modification within the artery wall that results in selective functional impairments. Plasma levels of this modified apoA-I form may provide insights into a pathophysiological process within the diseased artery wall. PMID:24558038

  6. Emerging Roles of Apolipoprotein E and Apolipoprotein A-I in the Pathogenesis and Treatment of Lung Disease.

    PubMed

    Yao, Xianglan; Gordon, Elizabeth M; Figueroa, Debbie M; Barochia, Amisha V; Levine, Stewart J

    2016-08-01

    Emerging roles are being recognized increasingly for apolipoproteins in the pathogenesis and treatment of lung diseases on the basis of their ability to suppress inflammation, oxidative stress, and tissue remodeling, and to promote adaptive immunity and host defense. Apolipoproteins, such as apolipoprotein E (apoE) and apolipoprotein A-I (apoA-I), are important components of lipoprotein particles that facilitate the transport of cholesterol, triglycerides, and phospholipids between plasma and cells. ApoE-containing lipoprotein particles are internalized into cells by low-density lipoprotein receptors (LDLRs), whereas apoA-I can interact with the ATP-binding cassette subfamily A member 1 (ABCA1) transporter to efflux cholesterol and phospholipids out of cells. ApoE and apoA-I also mediate receptor-independent effects, such as binding to and neutralizing LPS. Both apoE and apoA-I are expressed by lung cells, which allows apoE/LDLR- and apoA-I/ABCA1-dependent pathways to modulate normal lung health and the pathogenesis of respiratory diseases, including asthma, acute lung injury, cancer, emphysema, pulmonary fibrosis, and pulmonary hypertension. Data from human studies and research using experimental murine model systems have shown that both apoE and apoA-I pathways play primarily protective roles in lung biology and respiratory disease. Furthermore, apolipoprotein mimetic peptides, corresponding to the LDLR-binding domain of apoE or the class A amphipathic α-helical structure of apoA-I, have antiinflammatory and antioxidant effects that attenuate the severity of lung disease in murine models. Thus, the development of inhaled apolipoprotein mimetic peptides as a novel treatment paradigm could represent a significant advance for patients with respiratory disease who do not respond to current therapies.

  7. Emerging Roles of Apolipoprotein E and Apolipoprotein A-I in the Pathogenesis and Treatment of Lung Disease

    PubMed Central

    Yao, Xianglan; Gordon, Elizabeth M.; Figueroa, Debbie M.; Barochia, Amisha V.

    2016-01-01

    Emerging roles are being recognized increasingly for apolipoproteins in the pathogenesis and treatment of lung diseases on the basis of their ability to suppress inflammation, oxidative stress, and tissue remodeling, and to promote adaptive immunity and host defense. Apolipoproteins, such as apolipoprotein E (apoE) and apolipoprotein A-I (apoA-I), are important components of lipoprotein particles that facilitate the transport of cholesterol, triglycerides, and phospholipids between plasma and cells. ApoE-containing lipoprotein particles are internalized into cells by low-density lipoprotein receptors (LDLRs), whereas apoA-I can interact with the ATP-binding cassette subfamily A member 1 (ABCA1) transporter to efflux cholesterol and phospholipids out of cells. ApoE and apoA-I also mediate receptor-independent effects, such as binding to and neutralizing LPS. Both apoE and apoA-I are expressed by lung cells, which allows apoE/LDLR- and apoA-I/ABCA1-dependent pathways to modulate normal lung health and the pathogenesis of respiratory diseases, including asthma, acute lung injury, cancer, emphysema, pulmonary fibrosis, and pulmonary hypertension. Data from human studies and research using experimental murine model systems have shown that both apoE and apoA-I pathways play primarily protective roles in lung biology and respiratory disease. Furthermore, apolipoprotein mimetic peptides, corresponding to the LDLR-binding domain of apoE or the class A amphipathic α-helical structure of apoA-I, have antiinflammatory and antioxidant effects that attenuate the severity of lung disease in murine models. Thus, the development of inhaled apolipoprotein mimetic peptides as a novel treatment paradigm could represent a significant advance for patients with respiratory disease who do not respond to current therapies. PMID:27073971

  8. High-Throughput Analysis Identifying Drugs That Regulate Apolipoprotein A-I Synthesis.

    PubMed

    Haas, Michael J; Onstead-Haas, Luisa; Kurban, William; Shah, Harshit; Plazarte, Monica; Chamseddin, Ayham; Mooradian, Arshag D

    2017-07-25

    Apolipoprotein A-I (apo A-I) is the primary antiatherogenic protein in high-density lipoprotein (HDL). Despite the controversy as to the clinical effectiveness of raising HDL, the search is ongoing for safe and effective drugs that increase HDL and apo A-I levels. To identify novel compounds that can increase hepatic apo A-I production, two drug libraries were screened. The NIH clinical collection (NCC) and the NIH clinical collection 2 (NCC2) were purchased from Evotec (San Francisco, CA). The NCC library contains 446 compounds and the NCC2 library contains 281 compounds, all dissolved in dimethylsulfoxide at a concentration of 10 mM. Hepatoma-derived cells (HepG2) and primary hepatocytes in culture were treated with various compounds for 24 h and apo A-I in media samples was measured by enzyme immunoassay. Samples with significant changes in apo A-I concentrations were retested in independent experiments by Western blot analysis to confirm the immunoassay findings. Of a total of 727 compounds screened at a concentration of 50 μM, 15 compounds increased hepatic apo A-I production by 35%-54%, and 9 compounds lowered hepatic apo A-I concentrations in the culture media by 25%-52%. Future trials should explore the clinical effectiveness of these agents when standard doses of these drugs are used in humans.

  9. Hereditary apolipoprotein AI-associated renal amyloidosis: A diagnostic challenge.

    PubMed

    Samillán-Sosa, Kelly Del Rocío; Sención-Martínez, Gloria; Lopes-Martín, Vanessa; Martínez-González, Miguel Angel; Solé, Manel; Arostegui, Jose Luis; Mesa, Jose; García-Díaz, Juan de Dios; Rodríguez-Puyol, Diego; Martínez-Miguel, Patricia

    2015-01-01

    Hereditary renal amyloidosis is an autosomal dominant condition with considerable overlap with other amyloidosis types. Differential diagnosis is complicated, but is relevant for prognosis and treatment. We describe a patient with nephrotic syndrome and progressive renal failure, who had a mother with renal amiloidosis. Renal biopsy revealed amyloid deposits in glomerular space, with absence of light chains and protein AA. We suspected amyloidosis with fibrinogen A alpha chain deposits, which is the most frequent cause of hereditary amyloidosis in Europe, with a glomerular preferential affectation. However, the genetic study showed a novel mutation in apolipoprotein AI. On reviewing the biopsy of the patient's mother similar glomerular deposits were found, but there were significant deposits in the renal medulla as well, which is typical in APO AI amyloidosis. The diagnosis was confirmed by immunohistochemistry. Apo AI amyloidosis is characterized by slowly progressive renal disease and end-stage renal disease occurs aproximately 3 to 15 years from initial diagnosis. Renal transplantation offers an acceptable graft survival and in these patients with hepatorenal involvement simultaneous liver and kidney transplantation could be considered. Copyright © 2015 The Authors. Published by Elsevier España, S.L.U. All rights reserved.

  10. The promise of apolipoprotein A-I mimetics.

    PubMed

    Mendez, Armando J

    2010-04-01

    Synthetic high-density lipoprotein (HDL) and apolipoprotein (apo) A-I mimetic peptides emulate many of the atheroprotective biological functions attributed to HDL and can modify atherosclerotic disease processes. Administration of these agents as HDL replacement or modifying therapy has tremendous potential of providing new treatments for cardiovascular disease. Progress in the understanding of these agents is discussed in this review. Prospective, observational, and interventional studies have convincingly demonstrated that elevated serum levels of high-density lipoprotein-cholesterol (HDL-C) are associated with reduced risk for coronary heart disease (CHD). Although traditional pharmacological agents have shown modest utility in raising HDL levels and reducing CHD risk, use of HDL and apo A-I mimetics provides novel therapies to not only increase HDL levels, but to also influence HDL functionality. Evidence developed over the last several years has identified a number of pathways affected by synthetic HDL and apoA-I mimetic peptides, including enhancing reverse cholesterol transport and reducing oxidation and inflammation that directly influence the progression and regression of atherosclerotic disease. Clinical trials of relatively short-term synthetic HDL infusion into patients with CHD demonstrate beneficial effects. Use of apo A-I mimetic peptides could potentially overcome some of the limitations associated with use of the intact apo. Studies to establish the most efficacious peptides, optimal dosing regimens, and routes of administration are needed. Use of apo A-I mimetic peptides shows great promise as a therapeutic modality for HDL replacement and enhancing HDL function in treatment of patients with CHD.

  11. DNA inversion within the apolipoproteins AI/CIII/AIV-encoding gene cluster of certain patients with premature atherosclerosis

    SciTech Connect

    Karathanasis, S.K.; Ferris, E.; Haddad, I.A.

    1987-10-01

    The genes coding for apolipoproteins (apo) AI, CIII, and AIV, designated APOA1, APOC3, and APOA4, respectively, are closely linked and tandemly organized in the long arm of the human chromosome 11. A DNA rearrangement involving the genes encoding apoAI and apoCIII in certain patients with premature atherosclerosis has been associated with deficiency of both apoAI and apoCIII in the plasma of these patients. Structural characterization of the genes for apoAI and apoCIII in one of these patients indicates that this rearrangement consists of a DNA inversion containing portions of the 3' ends of the apoAI and apoCIII genes, including the DNA region between these genes. The breakpoints of this DNA inversion are located within the fourth exon of the apoAI gene and the first intron of the apoCIII gene. Thus, this DNA inversion results in reciprocal fusion of the apoAI and apoCIII gene transcriptional units. Expression of these gene fusions in cultured mammalian cells results in stable mRNA transcripts with sequences representing fusions of the apoAI and apoCIII mRNAs. These results indicate that absence of transcripts with correct apoAI and apoCIII mRNA sequences causes apoAI and apoCIII deficiency in the plasma of these patients and suggest that these apolipoproteins are involved in cholesterol homeostasis and protection against premature atherosclerosis.

  12. Apolipoproteins A-I, A-II and E in cholestatic liver disease.

    PubMed

    Florén, C H; Gustafson, A

    1985-04-01

    Apolipoproteins A-I, A-II and E were determined in the plasma of nine patients (five females, four males) with cholestatic liver disease (eight patients with primary biliary cirrhosis and one patient with sclerosing cholangitis). Plasma concentrations were measured by electroimmunoassay in the fasting state, postprandially after ingestion of either 100 g fat as whipping cream or a light mixed meal with or without addition of wheat fibre. Concentrations of apolipoproteins A-I and A-II were low in patients with cholestatic liver disease and A-I levels correlated inversely with the severity of liver disease as measured by bilirubin levels (r = -0.66). No changes in plasma apolipoprotein A-I, A-II or E concentrations occurred postprandially. There was an inverse correlation between plasma concentrations of apolipoproteins A-I and E (p less than 0.05, r = -0.68). A close relation existed between the ratio of apolipoprotein E to apolipoprotein A-I and plasma bile salt concentration (r = 0.80, p less than 0.01) and serum bilirubin (r = 0.76, p less than 0.01). This implies that in cholestatic liver disease apolipoprotein E and A-I levels reflect the degree of cholestasis.

  13. Ictalurus punctatus apolipoprotein A-I (ApoA1) mRNA, complete cds

    USDA-ARS?s Scientific Manuscript database

    The complete coding sequence of channel catfish apolipoprotein A-I is 777 bp in length, encoding 258 amino acids. The publishing of this coding sequence will also allow phylogenetic comparison between catfish ApoAI and ApoAI genes from other species. The availability of this complete coding sequence...

  14. Apolipoprotein AI Deficiency Inhibits Serum Opacity Factor Activity against Plasma High Density Lipoprotein via a Stabilization Mechanism

    PubMed Central

    Rosales, Corina; Patel, Niket; Gillard, Baiba K.; Yelamanchili, Dedipya; Yang, Yaliu; Courtney, Harry S.; Santos, Raul D.; Gotto, Antonio M.; Pownall, Henry J.

    2016-01-01

    The reaction of Streptococcal serum opacity factor (SOF) against plasma high-density lipoproteins (HDL) produces a large cholesteryl ester-rich microemulsion (CERM), a smaller neo HDL that is apolipoprotein (apo) AI-poor, and lipid-free apo AI. SOF is active vs. both human and mouse plasma HDL. In vivo injection of SOF into mice reduces plasma cholesterol ~40% in 3 hours while forming the same products observed in vitro, but at different ratios. Previous studies supported the hypothesis that labile apo AI is required for the SOF reaction vs. HDL. Here we further tested that hypothesis by studies of SOF against HDL from apo AI-null mice. When injected into apo AI-null mice, SOF reduced plasma cholesterol ~35% in three hours. The reaction of SOF vs. apo AI-null HDL in vitro produced a CERM and neo HDL, but no lipid-free apo. Moreover, according to the rate of CERM formation, the extent and rate of the SOF reaction vs. apo AI-null mouse HDL was less than that against wild-type (WT) mouse HDL. Chaotropic perturbation studies using guanidine hydrochloride showed that apo AI-null HDL was more stable than WT HDL. Human apo AI added to apo AI-null HDL was quantitatively incorporated, giving reconstituted HDL. Both SOF and guanidine hydrochloride displaced apo AI from the reconstituted HDL. These results support the conclusion that apo AI-null HDL is more stable than WT HDL because it lacks apo AI, a labile protein that is readily displaced by physico-chemical and biochemical perturbations. Thus, apo AI-null HDL is less SOF-reactive than WT HDL. The properties of apo AI-null HDL can be partially restored to those of WT HDL by the spontaneous incorporation of human apo AI. It remains to be determined what other HDL functions are affected by apo AI deletion. PMID:25790332

  15. Apolipoprotein AI deficiency inhibits serum opacity factor activity against plasma high density lipoprotein via a stabilization mechanism.

    PubMed

    Rosales, Corina; Patel, Niket; Gillard, Baiba K; Yelamanchili, Dedipya; Yang, Yaliu; Courtney, Harry S; Santos, Raul D; Gotto, Antonio M; Pownall, Henry J

    2015-04-14

    The reaction of Streptococcal serum opacity factor (SOF) against plasma high-density lipoproteins (HDL) produces a large cholesteryl ester-rich microemulsion (CERM), a smaller neo HDL that is apolipoprotein (apo) AI-poor, and lipid-free apo AI. SOF is active versus both human and mouse plasma HDL. In vivo injection of SOF into mice reduces plasma cholesterol ∼40% in 3 h while forming the same products observed in vitro, but at different ratios. Previous studies supported the hypothesis that labile apo AI is required for the SOF reaction vs HDL. Here we further tested that hypothesis by studies of SOF against HDL from apo AI-null mice. When injected into apo AI-null mice, SOF reduced plasma cholesterol ∼35% in 3 h. The reaction of SOF vs apo AI-null HDL in vitro produced a CERM and neo HDL, but no lipid-free apo. Moreover, according to the rate of CERM formation, the extent and rate of the SOF reaction versus apo AI-null mouse HDL were less than that against wild-type (WT) mouse HDL. Chaotropic perturbation studies using guanidine hydrochloride showed that apo AI-null HDL was more stable than WT HDL. Human apo AI added to apo AI-null HDL was quantitatively incorporated, giving reconstituted HDL. Both SOF and guanidine hydrochloride displaced apo AI from the reconstituted HDL. These results support the conclusion that apo AI-null HDL is more stable than WT HDL because it lacks apo AI, a labile protein that is readily displaced by physicochemical and biochemical perturbations. Thus, apo AI-null HDL is less SOF-reactive than WT HDL. The properties of apo AI-null HDL can be partially restored to those of WT HDL by the spontaneous incorporation of human apo AI. It remains to be determined what other HDL functions are affected by apo AI deletion.

  16. The Effects of Ginger on Fasting Blood Sugar, Hemoglobin A1c, Apolipoprotein B, Apolipoprotein A-I and Malondialdehyde in Type 2 Diabetic Patients

    PubMed Central

    Khandouzi, Nafiseh; Shidfar, Farzad; Rajab, Asadollah; Rahideh, Tayebeh; Hosseini, Payam; Mir Taheri, Mohsen

    2015-01-01

    Diabetes mellitus is the most common endocrine disorder, causes many complications such as micro- and macro-vascular diseases. Anti-diabetic, hypolipidemic and anti-oxidative properties of ginger have been noticed in several researches. The present study was conducted to investigate the effects of ginger on fasting blood sugar, Hemoglobin A1c, apolipoprotein B, apolipoprotein A-I, and malondialdehyde in type 2 diabetic patients. In a randomized, double-blind, placebo-controlled, clinical trial, a total of 41 type 2 diabetic patients randomly were assigned to ginger or placebo groups (22 in ginger group and 19 in control group), received 2 g/day of ginger powder supplement or lactose as placebo for 12 weeks. The serum concentrations of fasting blood sugar, Hemoglobin A1c, apolipoprotein B, apolipoprotein A-I and malondialdehyde were analyzed before and after the intervention. Ginger supplementation significantly reduced the levels of fasting blood sugar, hemoglobin A1c, apolipoprotein B, apolipoprotein B/apolipoprotein A-I and malondialdehyde in ginger group in comparison to baseline, as well as control group, while it increased the level of apolipoprotein A-I (p<0.05). It seems that oral administration of ginger powder supplement can improves fasting blood sugar, hemoglobin A1c, apolipoprotein B, apolipoprotein A-I, apolipoprotein B/apolipoprotein A-I and malondialdehyde in type 2 diabetic patients. So it may have a role in alleviating the risk of some chronic complications of diabetes. PMID:25561919

  17. The effects of ginger on fasting blood sugar, hemoglobin a1c, apolipoprotein B, apolipoprotein a-I and malondialdehyde in type 2 diabetic patients.

    PubMed

    Khandouzi, Nafiseh; Shidfar, Farzad; Rajab, Asadollah; Rahideh, Tayebeh; Hosseini, Payam; Mir Taheri, Mohsen

    2015-01-01

    Diabetes mellitus is the most common endocrine disorder, causes many complications such as micro- and macro-vascular diseases. Anti-diabetic, hypolipidemic and anti-oxidative properties of ginger have been noticed in several researches. The present study was conducted to investigate the effects of ginger on fasting blood sugar, Hemoglobin A1c, apolipoprotein B, apolipoprotein A-I, and malondialdehyde in type 2 diabetic patients. In a randomized, double-blind, placebo-controlled, clinical trial, a total of 41 type 2 diabetic patients randomly were assigned to ginger or placebo groups (22 in ginger group and 19 in control group), received 2 g/day of ginger powder supplement or lactose as placebo for 12 weeks. The serum concentrations of fasting blood sugar, Hemoglobin A1c, apolipoprotein B, apolipoprotein A-I and malondialdehyde were analyzed before and after the intervention. Ginger supplementation significantly reduced the levels of fasting blood sugar, hemoglobin A1c, apolipoprotein B, apolipoprotein B/apolipoprotein A-I and malondialdehyde in ginger group in comparison to baseline, as well as control group, while it increased the level of apolipoprotein A-I (p<0.05). It seems that oral administration of ginger powder supplement can improves fasting blood sugar, hemoglobin A1c, apolipoprotein B, apolipoprotein A-I, apolipoprotein B/apolipoprotein A-I and malondialdehyde in type 2 diabetic patients. So it may have a role in alleviating the risk of some chronic complications of diabetes.

  18. Identification of Apolipoprotein A-I as a Retinoic Acid-binding Protein in the Eye.

    PubMed

    Summers, Jody A; Harper, Angelica R; Feasley, Christa L; Van-Der-Wel, Hanke; Byrum, Jennifer N; Hermann, Marcela; West, Christopher M

    2016-09-02

    All-trans-retinoic acid may be an important molecular signal in the postnatal control of eye size. The goal of this study was to identify retinoic acid-binding proteins secreted by the choroid and sclera during visually guided ocular growth. Following photoaffinity labeling with all-trans-[11,12-(3)H]retinoic acid, the most abundant labeled protein detected in the conditioned medium of choroid or sclera had an apparent Mr of 27,000 Da. Following purification and mass spectrometry, the Mr 27,000 band was identified as apolipoprotein A-I. Affinity capture of the radioactive Mr 27,000 band by anti-chick apolipoprotein A-I antibodies confirmed its identity as apolipoprotein A-I. Photoaffinity labeling and fluorescence quenching experiments demonstrated that binding of retinoic acid to apolipoprotein A-I is 1) concentration-dependent, 2) selective for all-trans-retinoic acid, and 3) requires the presence of apolipoprotein A-I-associated lipids for retinoid binding. Expression of apolipoprotein A-I mRNA and protein synthesis were markedly up-regulated in choroids of chick eyes during the recovery from induced myopia, and apolipoprotein A-I mRNA was significantly increased in choroids following retinoic acid treatment. Together, these data suggest that apolipoprotein A-I may participate in a regulatory feedback mechanism with retinoic acid to control the action of retinoic acid on ocular targets during postnatal ocular growth. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Apolipoprotein A-I Q[-2]X causing isolated apolipoprotein A-I deficiency in a family with analphalipoproteinemia.

    PubMed Central

    Ng, D S; Leiter, L A; Vezina, C; Connelly, P W; Hegele, R A

    1994-01-01

    We report a Canadian kindred with a novel mutation in the apolipoprotein (apo) A-I gene causing analphalipoproteinemia. The 34-yr-old proband, product of a consanguineous marriage, had bilateral retinopathy, bilateral cataracts, spinocerebellar ataxia, and tendon xanthomata. High density lipoprotein cholesterol (HDL-C) was < 0.1 mM and apoA-I was undetectable. Genomic DNA sequencing of the proband's apoA-I gene identified a nonsense mutation at codon [-2], which we designate as Q[-2]X. This mutation causes a loss of endonuclease digestion sites for both BbvI and Fnu4HI. Genotyping identified four additional homozygotes, four heterozygotes, and two unaffected subjects among the first-degree relatives. Q[-2]X homozygosity causes a selective failure to produce any portion of mature apoA-I, resulting in very low plasma level of HDL. Heterozygosity results in approximately half-normal apoA-I and HDL. Gradient gel electrophoresis and differential electroimmunodiffusion assay revealed that the HDL particles of the homozygotes had peak Stokes diameter of 7.9 nm and contained apoA-II without apoA-I (Lp-AII). Heterozygotes had an additional fraction of HDL3-like particles. Two of the proband's affected sisters had documented premature coronary heart disease. This kindred, the third reported apoA-I gene mutation causing isolated complete apoA-I deficiency, appears to be at significantly increased risk for atherosclerosis. Images PMID:8282791

  20. Apolipoprotein A-I interactions with insulin secretion and production.

    PubMed

    Rye, Kerry-Anne; Barter, Philip J; Cochran, Blake J

    2016-02-01

    Human population studies have established that an elevated plasma high-density lipoprotein cholesterol (HDL-C) level is associated with a decreased risk of developing cardiovascular disease. In addition to having several potentially cardioprotective functions, HDLs and apolipoprotein (apo)A-I, the main HDL apolipoprotein, also have antidiabetic properties. Interventions that elevate plasma HDL-C and apoA-I levels improve glycemic control in people with type 2 diabetes mellitus by enhancing pancreatic β-cell function and increasing insulin sensitivity. This review is concerned with recent advances in understanding the mechanisms by which HDLs and apoA-I improve pancreatic β-cell function. HDLs and apoA-I increase insulin synthesis and secretion in pancreatic β cells. The underlying mechanism of this effect is similar to what has been reported for intestinally derived incretins, such as glucagon-like peptide-1 and glucose-dependent insulinotropic polypeptide, which both increase β-cell insulin secretion under high glucose conditions. This involves the activation of a heterotrimeric G protein Gαs subunit on the β-cell surface that leads to induction of a transmembrane adenylyl cyclase, increased intracellular cyclic adenosine monophosphate and Ca levels, and activation of protein kinase A. Protein kinase A increases insulin synthesis by excluding FoxO1 from the β-cell nucleus and derepressing transcription of the insulin gene.

  1. Molecules that mimic apolipoprotein A-I: potential agents for treating atherosclerosis.

    PubMed

    Leman, Luke J; Maryanoff, Bruce E; Ghadiri, M Reza

    2014-03-27

    Certain amphipathic α-helical peptides can functionally mimic many of the properties of full-length apolipoproteins, thereby offering an approach to modulate high-density lipoprotein (HDL) for combating atherosclerosis. In this Perspective, we summarize the key findings and advances over the past 25 years in the development of peptides that mimic apolipoproteins, especially apolipoprotein A-I (apoA-I). This assemblage of information provides a reasonably clear picture of the state of the art in the apolipoprotein mimetic field, an appreciation of the potential for such agents in pharmacotherapy, and a sense of the opportunities for optimizing the functional properties of HDL.

  2. Molecules that Mimic Apolipoprotein A-I: Potential Agents for Treating Atherosclerosis

    PubMed Central

    Leman, Luke J.; Maryanoff, Bruce E.; Ghadiri, M. Reza

    2013-01-01

    Certain amphipathic α-helical peptides can functionally mimic many of the properties of full-length apolipoproteins, thereby offering an approach to modulate high-density lipoprotein (HDL) for combating atherosclerosis. In this Perspective, we summarize the key findings and advances over the past 25 years in the development of peptides that mimic apolipoproteins, especially apolipoprotein A-I (apoA-I). This assemblage of information provides a reasonably clear picture of the state of the art in the apolipoprotein mimetic field, an appreciation of the potential for such agents in pharmacotherapy, and a sense of the opportunities for optimizing the functional properties of HDL. PMID:24168751

  3. Apolipoprotein AI and Transthyretin as Components of Amyloid Fibrils in a Kindred with apoAI Leu178His Amyloidosis

    PubMed Central

    de Sousa, Mónica Mendes; Vital, Claude; Ostler, Dominique; Fernandes, Rui; Pouget-Abadie, Jean; Carles, Dominique; Saraiva, Maria João

    2000-01-01

    We found a new C-terminal amyloidogenic variant of apolipoprotein AI (apoAI), Leu178His in a French kindred, associated with cardiac and larynx amyloidosis and skin lesions with onset during the fourth decade. This single-point mutation in exon 4 of the apoAI gene was detected by DNA sequencing of polymerase chain reaction amplified material and restriction fragment length polymorphism analysis in two siblings. Blood, larynx, and skin biopsies were available from one sibling. Anti-apoAI immunoblotting of isoelectric focusing of plasma showed a +1 alteration in the charge of the protein. Extraction of fibrils from the skin biopsy revealed both full-length and N-terminal fragments of apoAI and transthyretin (TTR). ApoAI and TTR co-localized in amyloid deposits as demonstrated by immunohistochemistry. The present report, together with the first recently described C-terminal amyloidogenic variant of apoAI, Arg173Pro, shows that amyloidogenicity of apoAI is not a feature exclusive to N-terminal variants. The most striking characteristic of amyloid fibrils in Leu178His is that wild-type TTR is co-localized with apoAI in the fibrils. We have previously determined that a fraction of plasma TTR circulates in plasma bound to high-density lipoprotein and that this interaction occurs through binding to apoAI. Therefore we hypothesize that nonmutated TTR might influence deposition of apoAI as amyloid. PMID:10854214

  4. Apolipoprotein AI and transthyretin as components of amyloid fibrils in a kindred with apoAI Leu178His amyloidosis.

    PubMed

    de Sousa, M M; Vital, C; Ostler, D; Fernandes, R; Pouget-Abadie, J; Carles, D; Saraiva, M J

    2000-06-01

    We found a new C-terminal amyloidogenic variant of apolipoprotein AI (apoAI), Leu178His in a French kindred, associated with cardiac and larynx amyloidosis and skin lesions with onset during the fourth decade. This single-point mutation in exon 4 of the apoAI gene was detected by DNA sequencing of polymerase chain reaction amplified material and restriction fragment length polymorphism analysis in two siblings. Blood, larynx, and skin biopsies were available from one sibling. Anti-apoAI immunoblotting of isoelectric focusing of plasma showed a +1 alteration in the charge of the protein. Extraction of fibrils from the skin biopsy revealed both full-length and N-terminal fragments of apoAI and transthyretin (TTR). ApoAI and TTR co-localized in amyloid deposits as demonstrated by immunohistochemistry. The present report, together with the first recently described C-terminal amyloidogenic variant of apoAI, Arg173Pro, shows that amyloidogenicity of apoAI is not a feature exclusive to N-terminal variants. The most striking characteristic of amyloid fibrils in Leu178His is that wild-type TTR is co-localized with apoAI in the fibrils. We have previously determined that a fraction of plasma TTR circulates in plasma bound to high-density lipoprotein and that this interaction occurs through binding to apoAI. Therefore we hypothesize that nonmutated TTR might influence deposition of apoAI as amyloid.

  5. Expression and recovery of biologically active recombinant Apolipoprotein AI(Milano) from transgenic safflower (Carthamus tinctorius) seeds.

    PubMed

    Nykiforuk, Cory L; Shen, Yin; Murray, Elizabeth W; Boothe, Joseph G; Busseuil, David; Rhéaume, Eric; Tardif, Jean-Claude; Reid, Alexandra; Moloney, Maurice M

    2011-02-01

    Apolipoprotein AI Milano (ApoAI(Milano) ) was expressed as a fusion protein in transgenic safflower seeds. High levels of expression corresponding to 7 g of ApoAI(Milano) per kilogram of seed have been identified in a line selected for commercialization. The ApoAI(Milano) fusion protein was extracted from seed using an oilbody-based process and matured in vitro prior to final purification. This yielded a Des-1,2-ApoAI(Milano) product which was confirmed by biochemical characterization including immunoreactivity against ApoAI antibodies, isoelectric point, N-terminal sequencing and electrospray mass spectrometry. Purified Des-1,2-ApoAI(Milano) readily associated with dimyristoylphosphatidylcholine in clearance assays comparable to Human ApoAI. Its biological activity was assessed by cholesterol efflux assays using Des-1,2-ApoAI(Milano) :1-palmitoyl-2-oleoyl phosphatidylcholine complexes in vitro and in vivo. This study has established that high levels of biologically functional ApoAI(Milano) can be produced using a plant-based expression system.

  6. Renal apolipoprotein A-I amyloidosis: a rare and usually ignored cause of hereditary tubulointerstitial nephritis.

    PubMed

    Gregorini, Gina; Izzi, Claudia; Obici, Laura; Tardanico, Regina; Röcken, Christoph; Viola, Battista Fabio; Capistrano, Mariano; Donadei, Simona; Biasi, Luciano; Scalvini, Tiziano; Merlini, Giampaolo; Scolari, Francesco

    2005-12-01

    Apolipoprotein A-I amyloidosis is a rare, late-onset, autosomal dominant condition characterized by systemic deposition of amyloid in tissues, the major clinical problems being related to renal, hepatic, and cardiac involvement. Described is the clinical and histologic picture of renal involvement as a result of apolipoprotein A-I amyloidosis in five families of Italian ancestry. In all of the affected family members, the disease was caused by the Leu75Pro heterozygous mutation in exon 4 of apolipoprotein A-I gene, as demonstrated by direct sequencing and RFLP analysis. Immunohistochemistry confirmed that amyloid deposits were specifically stained with an anti-apolipoprotein A-I antibody. The clinical phenotype was mainly characterized by a variable combination of kidney and liver disturbance. The occurrence of renal involvement seemed to be almost universal, although its severity varied greatly ranging from subclinical organ damage to overt, slowly progressive renal dysfunction. The renal presentation was consistent with a tubulointerstitial disease, as suggested by the findings of defective urine-concentrating capacity, moderate polyuria, negative urinalysis, and mild tubular proteinuria. Histology confirmed tubulointerstitial nephritis. Surprising, amyloid was restricted to nonglomerular regions and limited to the renal medulla. This location of apolipoprotein A-I amyloid differs sharply from other systemic amyloidoses that are mainly characterized by glomerular and vascular deposits. The tubulointerstitial nephritis as a result of hereditary apolipoprotein A-I amyloidosis is a rare disease and a challenging diagnosis to recognize. Patients who present with familial tubulointerstitial nephritis associated with liver disease require a high index of suspicion for apolipoprotein A-I amyloidosis.

  7. Immunolocalization of cubilin, megalin, apolipoprotein J, and apolipoprotein A-I in the uterus and oviduct.

    PubMed

    Argraves, W Scott; Morales, Carlos R

    2004-12-01

    Spermatozoa maturation and capacitation occurring in the male and female reproductive tracts, respectively, involves the remodeling of the spermatozoa plasma membrane. Apolipoprotein J (apoJ) and apolipoprotein A-I (apoA-I) have been implicated in the process of lipid exchange from the spermatozoa plasma membrane to epithelial cells lining the male reproductive tract. Evidence suggests that this process is mediated by the cooperative action of the endocytic lipoprotein receptors megalin and cubilin, which are expressed at the apical surface of absorptive epithelia in various tissues, including the efferent ducts and epididymis. Here, we investigated the possibility that these receptors and their lipid-binding ligands, apoJ and apoA-I, might function similarly in the female reproductive tract. We show that megalin and cubilin are expressed in the uterine epithelium at all stages of the estrous cycle, maximally during estrous and metestrous stages. In the oviduct, there is pronounced expression of both megalin and cubilin in the nonciliated cells of the proximal oviduct and epithelial cells of the distal oviduct, particularly during estrous and metestrous stages. In both uterine and oviduct epithelial cells, megalin and cubilin were located on the apical regions of the cells, consistent with a distribution at the cell surface and in endosomes. ApoJ and apoA-I were both detected in apical regions of uterine and oviduct epithelial cells. Secretory cells of the uterine glands were found to express apoJ and apoA-I suggesting that the glands are a site of synthesis for both proteins. In summary, our findings indicate that megalin and cubilin function within the female reproductive tract, possibly mediating uterine and oviduct epithelial cell endocytosis of apoJ/apoA-I-lipid complexes and thus playing a role in lipid efflux from the sperm plasma membrane, a major initiator of capacitation.

  8. Effect of urotensin II on apolipoprotein B100 and apolipoprotein A-I expression in HepG2 cell line

    PubMed Central

    Mohammadi, Abbas; Najar, Ahmad Gholamhoseinian; Khoshi, Amirhosein

    2014-01-01

    Background: Increased apolipoprotein B100 (apo B) and decreased apolipoprotein A-I (apo A-I) production are important risk factors in atherosclerosis. Urotensin II (UII), as the most potent vasoconstrictor in human, is related with hypertension and probably atherosclerosis. Because of the relationship between the hypertension and lipoprotein metabolism in atherosclerosis, the aim of this study was to test the effect of urotensin II on apo B and apo A-I expression in hepatic (HepG2) cell line. Materials and Methods: HepG2 cells were treated with 10, 50, 100, and 200 nmol/L of urotensin II (n = 6). Relative apo B and apo A-I messenger RNA (mRNA) levels in conditioned media, normalized to glyceraldehyde-3-phosphate dehydrogenase, were measured with quantitative real-time polymerase chain reaction method. In addition, apo B and apo A-I levels were also estimated and compared with the controls using the western blotting method. Data were analyzed statistically by ANOVA and non-parametric tests. Results: The apo B mRNA levels were not increased significantly following the treatment with UII. However, apo B protein levels were increased significantly after the treatment with urotensin II, especially at 100 and 200 nmol/L. The apo A-I mRNA and protein levels in conditioned media also were not significantly changed. However, there was a significant decrease in apo A-I mRNA and protein levels at 200 nM UII. Conclusions: UII might increase apo B at protein level probably through participating factors in its synthesis and/ or stability/degradation. In addition, UII may have decreasing effect at more than 200 nM concentrations on apo A-I. PMID:24600602

  9. Fructated apolipoprotein A-I exacerbates cellular senescence in human umbilical vein endothelial cells accompanied by impaired insulin secretion activity and embryo toxicity.

    PubMed

    Park, Ki-Hoon; Kim, Jae-Yong; Choi, Inho; Kim, Jae-Ryong; Won, Kyu Chang; Cho, Kyung-Hyun

    2016-08-01

    Glycation of apolipoproteins is a major feature of the production of dysfunctional high-density lipoprotein (HDL), which is associated with the incidence of several metabolic diseases such as coronary artery disease and diabetes. In this report, fructated apoA-I (fA-I) induced by fructose treatment showed a covalently multimerized band without cross-linking, and lysine residues were irreversibly modified to prevent crosslinking. Using pancreatic β-cells, insulin secretion was impaired by fA-I in the lipid-free and reconstituted HDL (rHDL) states, by up to 35%, and 40%, respectively, under hyperglycemic conditions (25 mmol/L glucose). Treatment of human umbilical vein endothelial cells (HUVECs) with fA-I and HDL from elderly patients caused a 1.8-fold and 1.5-fold increased cellular senescence, respectively, along with increased lysosomal enlargement. In the lipid-free and rHDL states, fA-I increased embryo death by 1.5-fold and 2.5-fold, respectively, along with the production of oxidized species. Furthermore, rHDL containing fA-I (fA-I-rHDL) showed a higher isoelectric point (pI, approximately 8.5), whereas rHDL containing nA-I (nA-I-rHDL) showed a narrow band range with lower pI (around 8.0) as well as a much smaller particle size than that of nA-I-rHDL. In conclusion, fructose-mediated apoA-I fructation resulted in the severe loss of several beneficial functions of apoA-I and HDL, including anti-senescence and insulin secretion activities, accompanied with increased susceptibility to protein degradation and structural modification.

  10. Bioinformatic Analysis of Plasma Apolipoproteins A-I and A-II Revealed Unique Features of A-I/A-II HDL Particles in Human Plasma

    PubMed Central

    Kido, Toshimi; Kurata, Hideaki; Kondo, Kazuo; Itakura, Hiroshige; Okazaki, Mitsuyo; Urata, Takeyoshi; Yokoyama, Shinji

    2016-01-01

    Plasma concentration of apoA-I, apoA-II and apoA-II-unassociated apoA-I was analyzed in 314 Japanese subjects (177 males and 137 females), including one (male) homozygote and 37 (20 males and 17 females) heterozygotes of genetic CETP deficiency. ApoA-I unassociated with apoA-II markedly and linearly increased with HDL-cholesterol, while apoA-II increased only very slightly and the ratio of apoA-II-associated apoA-I to apoA-II stayed constant at 2 in molar ratio throughout the increase of HDL-cholesterol, among the wild type and heterozygous CETP deficiency. Thus, overall HDL concentration almost exclusively depends on HDL with apoA-I without apoA-II (LpAI) while concentration of HDL containing apoA-I and apoA-II (LpAI:AII) is constant having a fixed molar ratio of 2 : 1 regardless of total HDL and apoA-I concentration. Distribution of apoA-I between LpAI and LpAI:AII is consistent with a model of statistical partitioning regardless of sex and CETP genotype. The analysis also indicated that LpA-I accommodates on average 4 apoA-I molecules and has a clearance rate indistinguishable from LpAI:AII. Independent evidence indicated LpAI:A-II has a diameter 20% smaller than LpAI, consistent with a model having two apoA-I and one apoA-II. The functional contribution of these particles is to be investigated. PMID:27526664

  11. Heparin promotes fibril formation by the N-terminal fragment of amyloidogenic apolipoprotein A-I.

    PubMed

    Mikawa, Shiho; Mizuguchi, Chiharu; Nishitsuji, Kazuchika; Baba, Teruhiko; Shigenaga, Akira; Shimanouchi, Toshinori; Sakashita, Naomi; Otaka, Akira; Akaji, Kenichi; Saito, Hiroyuki

    2016-10-01

    Glycosaminoglycans are known to be associated with extracellular amyloid deposits of various amyloidogenic proteins. In this study, we found that the glycosaminoglycan heparin greatly accelerates the elongation step in fibril formation by the N-terminal 1-83 fragment of human apolipoprotein A-I (apoA-I), especially in the amyloidogenic W50R variant. Using fragment peptides, we demonstrate that heparin significantly promotes β-transition and fibril formation of the highly amyloidogenic region spanning residues 44-65 and colocalizes with fibrils formed by the W50R variant. These results suggest the possible role of glycosaminoglycans in fibril formation by amyloidogenic apoA-I variants. © 2016 Federation of European Biochemical Societies.

  12. Learning from Synthetic Models of Extracellular Matrix; Differential Binding of Wild Type and Amyloidogenic Human Apolipoprotein A-I to Hydrogels Formed from Molecules Having Charges Similar to Those Found in Natural GAGs.

    PubMed

    Rosú, Silvana A; Toledo, Leandro; Urbano, Bruno F; Sanchez, Susana A; Calabrese, Graciela C; Tricerri, M Alejandra

    2017-08-01

    Among other components of the extracellular matrix (ECM), glycoproteins and glycosaminoglycans (GAGs) have been strongly associated to the retention or misfolding of different proteins inducing the formation of deposits in amyloid diseases. The composition of these molecules is highly diverse and a key issue seems to be the equilibrium between physiological and pathological events. In order to have a model in which the composition of the matrix could be finely controlled, we designed and synthesized crosslinked hydrophilic polymers, the so-called hydrogels varying the amounts of negative charges and hydroxyl groups that are prevalent in GAGs. We checked and compared by fluorescence techniques the binding of human apolipoprotein A-I and a natural mutant involved in amyloidosis to the hydrogel scaffolds. Our results indicate that both proteins are highly retained as long as the negative charge increases, and in addition it was shown that the mutant is more retained than the Wt, indicating that the retention of specific proteins in the ECM could be part of the pathogenicity. These results show the importance of the use of these polymers as a model to get deep insight into the studies of proteins within macromolecules.

  13. Novel therapies to increase apolipoprotein AI and HDL for the treatment of atherosclerosis.

    PubMed

    Wong, Norman Cw

    2007-09-01

    Apolipoprotein AI (apoAI) is the major protein component of HDL, and thus has an important role in the treatment of atherosclerosis. This review summarizes the various approaches being examined for raising levels of apoAI/HDL, including increasing the synthesis of apoAI and altering the metabolism of HDL. In addition, the currently available drugs used to increase apoAI/HDL are discussed, with a focus on the potential sites of action of these drugs on HDL metabolism. The outcome of further investigational studies into this field should provide effective therapies to increase apoAI/HDL levels and thus be of use in the treatment of cardiovascular disease.

  14. Chromatofocusing of human high density lipoproteins and isolation of lipoproteins A and A-I.

    PubMed

    Nestruck, A C; Niedmann, P D; Wieland, H; Seidel, D

    1983-08-29

    Using chromatofocusing, a column chromatography method with an internally generated pH gradient and focusing effects, human plasma high density lipoproteins (HDL) were fractionated into six subclasses within an interval of less than 1 pH unit (pH 5.1-4.2). All fractions floated in the ultracentrifuge at density = 1.21 g X ml-1, retained a typical HDL electron micrographic morphology and as a single band, alpha-migration on agarose electrophoresis. Compositional analysis of the subclasses revealed an inverse relationship between cholesterol ester and cholesterol on a molar basis. Distinct differences in the distribution of the apolipoproteins between the fractions were found. Two of the subclasses contained only apolipoprotein A-I and were therefore considered to be two forms of the lipid-combined form of apolipoprotein A-I, i.e., lipoprotein A-I. One subclass contained only apolipoproteins A-I + A-II and was, therefore, lipoprotein A. One subclass contained apolipoproteins A-I + A-II + D, and the two remaining contained additionally apolipoproteins C and E. Lipoprotein A-I was also demonstrated after immunoabsorption of apolipoprotein A-II-containing lipoproteins from whole serum. It is suggested that this method, which allows the fractionation of HDL into subclasses with distinct differences in apolipoprotein composition, offers new avenues for the study of the structural and metabolic heterogeneity of HDL.

  15. Apolipoprotein A-I Limits the Negative Effect of Tumor Necrosis Factor on Lymphangiogenesis.

    PubMed

    Bisoendial, Radjesh; Tabet, Fatiha; Tak, Paul P; Petrides, Francine; Cuesta Torres, Luisa F; Hou, Liming; Cook, Adam; Barter, Philip J; Weninger, Wolfgang; Rye, Kerry-Anne

    2015-11-01

    Lymphatic endothelial dysfunction underlies the pathogenesis of many chronic inflammatory disorders. The proinflammatory cytokine tumor necrosis factor (TNF) is known for its role in disrupting the function of the lymphatic vasculature. This study investigates the ability of apolipoprotein (apo) A-I, the principal apolipoprotein of high-density lipoproteins, to preserve the normal function of lymphatic endothelial cells treated with TNF. TNF decreased the ability of lymphatic endothelial cells to form tube-like structures. Preincubation of lymphatic endothelial cells with apoA-I attenuated the TNF-mediated inhibition of tube formation in a concentration-dependent manner. In addition, apoA-I reversed the TNF-mediated suppression of lymphatic endothelial cell migration and lymphatic outgrowth in thoracic duct rings. ApoA-I also abrogated the negative effect of TNF on lymphatic neovascularization in an ATP-binding cassette transporter A1-dependent manner. At the molecular level, this involved downregulation of TNF receptor-1 and the conservation of prospero-related homeobox gene-1 expression, a master regulator of lymphangiogenesis. ApoA-I also re-established the normal phenotype of the lymphatic network in the diaphragms of human TNF transgenic mice. ApoA-I restores the neovascularization capacity of the lymphatic system during TNF-mediated inflammation. This study provides a proof-of-concept that high-density lipoprotein-based therapeutic strategies may attenuate chronic inflammation via its action on lymphatic vasculature. © 2015 American Heart Association, Inc.

  16. Characterization of high density lipoprotein particles in familial apolipoprotein A-I deficiency

    USDA-ARS?s Scientific Manuscript database

    Our aim was to characterize HDL subspecies and fat-soluble vitamin levels in a kindred with familial apolipoprotein A-I (apoA-I) deficiency. Sequencing of the APOA1 gene revealed a nonsense mutation at codon 22, Q[22]X, with two documented homozygotes, eight heterozygotes, and two normal subjects in...

  17. Apolipoprotein A-I: the dual face of a protein.

    PubMed

    Arciello, Angela; Piccoli, Renata; Monti, Daria Maria

    2016-12-01

    Conformational plasticity and flexibility are key structural features of ApoAI in lipid metabolism. Amyloidogenic single point mutations, associated with incurable familial amyloidosis with fibril deposition in peripheral organs, may have a dramatic impact on the structural and functional features of ApoAI. Here, the consistent body of data on ApoAI variants has been reviewed, with the aim of highlighting the hallmarks of the pathology. In accordance with our observations, as well as that of others, we propose a model that accounts for the alteration of the delicate balance between lipid-free/lipid-bound dynamic states which is based on monomer-to-dimer interconversion via domain swapping. © 2016 Federation of European Biochemical Societies.

  18. Apolipoprotein A-I mimetic peptides inhibit expression and activity of hypoxia-inducible factor-1α in human ovarian cancer cell lines and a mouse ovarian cancer model.

    PubMed

    Gao, Feng; Chattopadhyay, Arnab; Navab, Mohamad; Grijalva, Victor; Su, Feng; Fogelman, Alan M; Reddy, Srinivasa T; Farias-Eisner, Robin

    2012-08-01

    Our previous results demonstrated that the apolipoprotein A-I (apoA-I) mimetic peptides L-4F and L-5F inhibit vascular endothelial growth factor production and tumor angiogenesis. The present study was designed to test whether apoA-I mimetic peptides inhibit the expression and activity of hypoxia-inducible factor-1α (HIF-1α), which plays a critical role in the production of angiogenic factors and angiogenesis. Immunohistochemistry staining was used to examine the expression of HIF-1α in tumor tissues. Immunoblotting, real-time polymerase chain reaction, immunofluorescence, and luciferase activity assays were used to determine the expression and activity of HIF-1α in human ovarian cancer cell lines. Immunohistochemistry staining demonstrated that L-4F treatment dramatically decreased HIF-1α expression in mouse ovarian tumor tissues. L-4F inhibited the expression and activity of HIF-1α induced by low oxygen concentration, cobalt chloride (CoCl(2), a hypoxia-mimic compound), lysophosphatidic acid, and insulin in two human ovarian cancer cell lines, OV2008 and CAOV-3. L-4F had no effect on the insulin-induced phosphorylation of Akt, but inhibited the activation of extracellular signal-regulated kinase and p70s6 kinase, leading to the inhibition of HIF-1α synthesis. Pretreatment with L-4F dramatically accelerated the proteasome-dependent protein degradation of HIF-1α in both insulin- and CoCl(2)-treated cells. The inhibitory effect of L-4F on HIF-1α expression is in part mediated by the reactive oxygen species-scavenging effect of L-4F. ApoA-I mimetic peptides inhibit the expression and activity of HIF-1α in both in vivo and in vitro models, suggesting the inhibition of HIF-1α may be a critical mechanism responsible for the suppression of tumor progression by apoA-I mimetic peptides.

  19. Inhibition of apolipoprotein A-I gene expression by obesity-associated endocannabinoids.

    PubMed

    Haas, Michael J; Mazza, Angela D; Wong, Norman C W; Mooradian, Arshag D

    2012-04-01

    Obesity is associated with increased serum endocannabinoid (EC) levels and decreased high-density lipoprotein cholesterol (HDLc). Apolipoprotein A-I (apo A-I), the primary protein component of HDL is expressed primarily in the liver and small intestine. To determine whether ECs regulate apo A-I gene expression directly, the effect of the obesity-associated ECs anandamide and 2-arachidonylglycerol on apo A-I gene expression was examined in the hepatocyte cell line HepG2 and the intestinal cell line Caco-2. Apo A-I protein secretion was suppressed nearly 50% by anandamide and 2-arachidonoylglycerol in a dose-dependent manner in both cell lines. Anandamide treatment suppressed both apo A-I mRNA and apo A-I gene promoter activity in both cell lines. Studies using apo A-I promoter deletion constructs indicated that repression of apo A-I promoter activity by anandamide requires a previously identified nuclear receptor binding site designated as site A. Furthermore, anandamide-treatment inhibited protein-DNA complex formation with the site A probe. Exogenous over expression of cannabinoid receptor 1 (CBR1) in HepG2 cells suppressed apo A-I promoter activity, while in Caco-2 cells, exogenous expression of both CBR1 and CBR2 could repress apo A-I promoter activity. The suppressive effect of anandamide on apo A-I promoter activity in Hep G2 cells could be inhibited by CBR1 antagonist AM251 but not by AM630, a selective and potent CBR2 inhibitor. These results indicate that ECs directly suppress apo A-I gene expression in both hepatocytes and intestinal cells, contributing to the decrease in serum HDLc in obese individuals.

  20. Insulin-Mediated Downregulation of Apolipoprotein A-I Gene in Human Hepatoma Cell Line HepG2: The Role of Interaction Between FOXO1 and LXRβ Transcription Factors.

    PubMed

    Shavva, Vladimir S; Bogomolova, Alexandra M; Nikitin, Artemy A; Dizhe, Ella B; Tanyanskiy, Dmitry A; Efremov, Alexander M; Oleinikova, Galina N; Perevozchikov, Andrej P; Orlov, Sergey V

    2017-02-01

    Apolipoprotein A-I (ApoA-I) is a key component of high density lipoproteins which possess anti-atherosclerotic and anti-inflammatory properties. Insulin is a crucial mediator of the glucose and lipid metabolism that has been implicated in atherosclerotic and inflammatory processes. Important mediators of insulin signaling such as Liver X Receptors (LXRs) and Forkhead Box A2 (FOXA2) are known to regulate apoA-I expression in liver. Forkhead Box O1 (FOXO1) is a well-known target of insulin signaling and a key mediator of oxidative stress response. Low doses of insulin were shown to activate apoA-I expression in human hepatoma HepG2 cells. However, the detailed mechanisms for these processes are still unknown. We studied the possible involvement of FOXO1, FOXA2, LXRα, and LXRβ transcription factors in the insulin-mediated regulation of apoA-I expression. Treatment of HepG2 cells with high doses of insulin (48 h, 100 nM) suppresses apoA-I gene expression. siRNAs against FOXO1, FOXA2, LXRβ, or LXRα abrogated this effect. FOXO1 forms a complex with LXRβ and insulin treatment impairs FOXO1/LXRβ complex binding to hepatic enhancer and triggers its nuclear export. Insulin as well as LXR ligand TO901317 enhance the interaction between FOXA2, LXRα, and hepatic enhancer. These data suggest that high doses of insulin downregulate apoA-I gene expression in HepG2 cells through redistribution of FOXO1/LXRβ complex, FOXA2, and LXRα on hepatic enhancer of apoA-I gene. J. Cell. Biochem. 118: 382-396, 2017. © 2016 Wiley Periodicals, Inc.

  1. Conformational analysis of apolipoprotein A-I and E-3 based on primary sequence and circular dichroism.

    PubMed Central

    Nolte, R T; Atkinson, D

    1992-01-01

    The primary and secondary structure of human plasma apolipoprotein A-I and apolipoprotein E-3 have been analyzed to further our understanding of the secondary and tertiary conformation of these proteins and the structure and function of plasma lipoprotein particles. The methods used to analyze the primary sequence of these proteins used computer programs: (a) to identify repeated patterns within these proteins on the basis of conservative substitutions and similarities within the physicochemical properties of each residue; (b) for local averaging, hydrophobic moment, and Fourier analysis of the physicochemical properties; and (c) for secondary structure prediction of each protein carried out using homology, statistical, and information theory based methods. Circular dichroism was used to study purified lipid-protein complexes of each protein and quantitate the secondary structure in a lipid environment. The data from these analyses were integrated into a single secondary structure prediction to derive a model of each protein. The sequence homology within apolipoproteins A-I, E-3, and A-IV is used to derive a consensus sequence for two 11 amino acid repeating sequences in this family of proteins. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 8 PMID:1477274

  2. The Effect of Aerobic Exercise on Total Cholesterol, High-Density Lipoprotein, Apolipoprotein B, Apolipoprotein A-I, and Percent Body Fat in Adolescent Females.

    ERIC Educational Resources Information Center

    Lungo, Diane; And Others

    The effect of aerobic exercise on total cholesterol (TC), high-density lipoprotein (HDL), apolipoprotein B (Apo B), apolioprotein A-I (Apo A-I), and percent body fat in adolescent females was studied. The control subjects (n=86) were volunteers who had completed a physical education class at least six months prior to the commencement of the study,…

  3. The Effect of Aerobic Exercise on Total Cholesterol, High-Density Lipoprotein, Apolipoprotein B, Apolipoprotein A-I, and Percent Body Fat in Adolescent Females.

    ERIC Educational Resources Information Center

    Lungo, Diane; And Others

    The effect of aerobic exercise on total cholesterol (TC), high-density lipoprotein (HDL), apolipoprotein B (Apo B), apolioprotein A-I (Apo A-I), and percent body fat in adolescent females was studied. The control subjects (n=86) were volunteers who had completed a physical education class at least six months prior to the commencement of the study,…

  4. Induction of apolipoprotein A-I gene expression by black seed (Nigella sativa) extracts.

    PubMed

    Haas, Michael J; Onstead-Haas, Luisa M; Naem, Emad; Wong, Norman C W; Mooradian, Arshag D

    2014-09-01

    Black seed [Nigella sativa L. (Ranunculaceae)] has been shown in animal models to lower serum cholesterol levels. In order to determine if extracts from black seed have any effects on high-density lipoprotein (HDL), we characterized the effects of black seed extract on apolipoprotein A-I (apo A-I) gene expression, the primary protein component of HDL. Hepatocytes (HepG2) and intestinal cells (Caco-2) were treated with black seed extracts, and Apo A-I, peroxisome proliferator-activated receptor α (PPARα), and retinoid-x-receptor α (RXRα) were measured by Western blot analysis. Apo A-I mRNA levels were measured by quantitative real-time polymerase chain reaction and apo A-I gene transcription was measured by transient transfection of apo A-I reporter plasmids. Extracts from black seeds significantly increased hepatic and intestinal apo A-I secretion, as well as apo A-I mRNA and gene promoter activity. This effect required a PPARα binding site in the apo A-I gene promoter. Treatment of the extract with either heat or trypsin had no effect on its ability to induce apo A-I secretion. Treatment with black seed extract induced PPARα expression 9-fold and RXRα expression 2.5-fold. Furthermore, the addition of PPARα siRNA but not a control siRNA prevented some but not all the positive effects of black seed on apo A-I secretion. Black seed extract is a potent inducer of apo A-I gene expression, presumably by enhancing PPARα/RXRα expression. We conclude that black seed may have beneficial effects in treating dyslipidemia and coronary heart disease.

  5. Reaction of discoidal complexes of apolipoprotein A-I and various phosphatidylcholines with lecithin cholesterol acyltransferase. Interfacial effects.

    PubMed

    Jonas, A; Zorich, N L; Kézdy, K E; Trick, W E

    1987-03-25

    Complexes of phospholipids-apolipoprotein A-I-cholesterol, containing various bulk phosphatidylcholines or a matrix of the ether analog of 1-palmitoyl 2-oleoyl phosphatidylcholine including test phosphatidylcholines were used as substrates for human lecithin-cholesterol acyltransferase. The enzymatic reaction rates for both series of complexes were determined as a function of temperature, particle concentration, neutral salt concentration, and the type of anion present in solution. The kinetic results support the hypothesis that phospholipids, in discoidal complexes, modulate the reaction rates by molecular effects at the active site, but also by interfacial effects on the interaction of the enzyme with the particles. The relevant interfacial parameters are the lipid packing at the interface and the structure of apolipoprotein A-I.

  6. Apolipoprotein A-I and its mimetics for the treatment of atherosclerosis

    PubMed Central

    Smith, Jonathan D

    2011-01-01

    Although statin treatment leads consistently to a reduction in major adverse coronary events and death in clinical trials, approximately 60 to 70% residual risk of these outcomes still remains. One frontier of investigational drug research is treatment to increase HDL, the ‘good cholesterol’ that is associated with a reduced risk of coronary artery disease. HDL and its major protein apolipoprotein A-I (apoAI) are protective against atherosclerosis through several mechanisms, including the ability to mediate reverse cholesterol transport. This review focuses on the preclinical and clinical findings for two types of therapies for the treatment of atherosclerosis: apoAI-containing compounds and apoAI mimetic peptides. Both of these therapies have excellent potential to be useful clinically to promote atherosclerosis regression and stabilize existing plaques, but significant hurdles must be overcome in order to develop these approaches into safe and effective therapies. PMID:20730693

  7. Regulation of the promoter of rat apolipoprotein A-I gene in cultured cells

    SciTech Connect

    Chao, Y.; Pan, T.; Wu, T.; Hao, Q.; Yamin, T.; Kroon, P.A.

    1987-05-01

    In order to study the regulation of the promoter of apolipoprotein (apo) A-I gene, they joined the 5' end of rat apo A-I gene (1.9 Kb) to the coding region of bacterial chloramphenicol acetyltransferase (CAT) gene. The chimeric gene produced high levels of CAT activity in both mouse L cells and Hep G2 cells in transient expression assays. Ethanol increased the levels of rat apo A-I promoter activity in both cells. However, dexamethasone increased rat apo A-I promoter activity only in Hep G2 cells. Similar results were obtained in stable expression cell lines. Nucleotide deletion experiments showed DNA sequences between -149 and -469 base pairs upstream from the rat apo A-I transcription site are required for the high level of expression and that the regulatory sequences are located further upstream. These data demonstrated that the 5' end of rat apo A-I gene contains sequences which are responsible for the regulation of apo A-I expression by ethanol and dexamethasone and that the expression and regulation of rat apo A-I promoter are cell specific.

  8. Effects of dietary maritime pine (Pinus pinaster)-seed oil on high-density lipoprotein levels and in vitro cholesterol efflux in mice expressing human apolipoprotein A-I.

    PubMed

    Asset, G; Leroy, A; Bauge, E; Wolff, R L; Fruchart, J C; Dallongeville, J

    2000-09-01

    Maritime pine (Pinus pinaster)-seed oil contains two Delta5 unsaturated polymethylene interrupted fatty acids (all cis-5,9, 12-18:3 and all cis-5,11,14-20:3 acids) one of which resembles eicosapentaenoic acid. The goal of the present study was to test whether maritime pine-seed oil consumption affects HDL and apolipoprotein (Apo) A-I levels as well as the ability of serum to promote efflux of cholesterol from cultured cells. To this end, wild type (WT) non-transgenic mice and transgenic mice expressing human ApoA-I (HuA-ITg) were fed on isoenergetic diet containing either 200 g maritime pine-seed oil/kg or 200 g lard/kg for 2 weeks. WT and HuA-ITg mice fed maritime pine-seed oil had lower cholesterol, HDL-cholesterol, LDL-cholesterol and HuA-ITg mice had lower human ApoA-I than those fed lard. The differences in cholesterol (P < 0.0001) and HDL-cholesterol (P < 0.003) levels between mice fed on the two diets were more pronounced in the HuA-ITg than in the WT mice. The ability of HuA-ITg serum to promote cholesterol efflux in cultured cells was greater (P < 0.008) than that of WT animals. However, the maritime pine-seed oil diet was associated with lower (P < 0.005) in vitro cholesterol efflux ability than the lard diet in both mice genotypes. This suggests a negative effect of the maritime pine-seed oil on reverse cholesterol transport. Cholesterol efflux was correlated with serum free or esterified cholesterol and phospholipid levels. The slope of the regression line was smaller in the HuA-ITg than in the WT mice indicating that overexpression of human ApoA-I reduces the negative impact of maritime pine-seed oil on cholesterol efflux. In conclusion, maritime pine-seed oil diet lowers HDL-cholesterol and diminishes in vitro cholesterol efflux. This potentially detrimental effect is attenuated by overexpression of human ApoA-I in mice.

  9. Apolipoprotein A-I mutant proteins having cysteine substitutions and polynucleotides encoding same

    DOEpatents

    Oda, Michael N.; Forte, Trudy M.

    2007-05-29

    Functional Apolipoprotein A-I mutant proteins, having one or more cysteine substitutions and polynucleotides encoding same, can be used to modulate paraoxonase's arylesterase activity. These ApoA-I mutant proteins can be used as therapeutic agents to combat cardiovascular disease, atherosclerosis, acute phase response and other inflammatory related diseases. The invention also includes modifications and optimizations of the ApoA-I nucleotide sequence for purposes of increasing protein expression and optimization.

  10. Apolipoprotein A-I metabolism in cynomolgus monkey. Identification and characterization of beta-migrating pools

    SciTech Connect

    Melchior, G.W.; Castle, C.K.

    1989-07-01

    Fresh plasma from control (C) and hypercholesterolemic (HC) cynomolgus monkeys was analyzed by agarose electrophoresis-immunoblotting with antibody to cynomolgus monkey apolipoprotein (apo) A-I. Two bands were evident on the autoradiogram: an alpha-migrating band (high density lipoprotein) and a beta-migrating band that comigrated exactly with cynomolgus monkey low density lipoprotein (LDL). The presence of beta-migrating apo A-I in the plasma of these monkeys was confirmed by Geon-Pevikon preparative electrophoresis, crossed immunoelectrophoresis, and isotope dilution studies in which radiolabeled apo A-I was found to equilibrate also with alpha- and beta-migrating pools of apo A-I in the plasma. Subfractionation of C and HC plasma by agarose column chromatography (Bio-Gel A-0.5M and A-15M) followed by agarose electrophoresis-immunoblotting indicated that the beta-migrating apo A-I in C was relatively homogeneous and eluted with proteins of Mr approximately 50 kD (apo A-I(50 kD)), whereas two beta-migrating fractions were identified in HC, one that eluted with the 50-kD proteins, and the other that eluted in the LDL Mr range (apo A-I(LDL)). The apo A-I(LDL) was precipitated by antibody to cynomolgus monkey apo B. The apo A-I(50 kD) accounted for 5 +/- 1% (mean +/- SD) of the plasma apo A-I in C plasma, and 15 +/- 7% in HC plasma. No apo A-I(LDL) was detected in C plasma, but that fraction accounted for 9 +/- 7% of the apo A-I in HC plasma. These data establish the presence of multiple pools of apo A-I in the cynomolgus monkey, which must be taken into consideration in any comprehensive model of apo A-I metabolism in this species.

  11. Iowa Mutant Apolipoprotein A-I (ApoA-IIowa) Fibrils Target Lysosomes

    PubMed Central

    Kameyama, Hirokazu; Nakajima, Hiroyuki; Nishitsuji, Kazuchika; Mikawa, Shiho; Uchimura, Kenji; Kobayashi, Norihiro; Okuhira, Keiichiro; Saito, Hiroyuki; Sakashita, Naomi

    2016-01-01

    The single amino acid mutation G26R in human apolipoprotein A-I (apoA-IIowa) is the first mutation that was associated with familial AApoA1 amyloidosis. The N-terminal fragments (amino acid residues 1–83) of apoA-I containing this mutation deposit as amyloid fibrils in patients’ tissues and organs, but the mechanisms of cellular degradation and cytotoxicity have not yet been clarified. In this study, we demonstrated degradation of apoA-IIowa fibrils via the autophagy-lysosomal pathway in human embryonic kidney 293 cells. ApoA-IIowa fibrils induced an increase in lysosomal pH and the cytosolic release of the toxic lysosomal protease cathepsin B. The mitochondrial dysfunction caused by apoA-IIowa fibrils depended on cathepsin B and was ameliorated by increasing the degradation of apoA-IIowa fibrils. Thus, although apoA-IIowa fibril transport to lysosomes and fibril degradation in lysosomes may have occurred, the presence of an excess number of apoA-IIowa fibrils, more than the lysosomes could degrade, may be detrimental to cells. Our results thus provide evidence that the target of apoA-IIowa fibrils is lysosomes, and we thereby gained a novel insight into the mechanism of AApoA1 amyloidosis. PMID:27464946

  12. Apolipoprotein AI could be a significant determinant of epithelial integrity in rainbow trout gill cell cultures: a study in functional proteomics.

    PubMed

    Smith, Richard W; Wood, Chris M; Cash, Phil; Diao, Linda; Pärt, Peter

    2005-05-20

    The freshwater fish gill forms a barrier against an external hypotonic environment. By culturing rainbow trout gill cells on permeable supports, as intact epithelia, this study investigates barrier property mechanisms. Under symmetrical conditions the apical and basolateral epithelial surfaces contact cell culture media. Replacing apical media with water, to generate asymmetrical conditions (i.e. the situation encountered by the freshwater gill), rapidly increases transepithelial resistance (TER). Proteomic analysis revealed that this is associated with enhanced expression of pre-apolipoprotein AI (pre-apoAI). To test the physiological relevance, gill cells were treated with a dose of 50 microg ml(-1) human apolipoprotein (apoAI). This was found to elevate TER in those epithelia which displayed a lower TER prior to apoAI treatment. These results demonstrate the action of apoAI and provide evidence that the rainbow trout gill may be a site of apoAI synthesis. TER does not differentiate between the trans-cellular (via the cell membrane) and para-cellular (via intercellular tight junctions) pathways. However, despite the apoAI-induced changes in TER, para-cellular permeability (measured by polyethylene glycol efflux) remained unaltered suggesting apoAI specifically reduces trans-cellular permeability. This investigation combines proteomics with functional measurements to show how a proteome change may be associated with freshwater gill function.

  13. Defective removal of cellular cholesterol and phospholipids by apolipoprotein A-I in Tangier Disease.

    PubMed Central

    Francis, G A; Knopp, R H; Oram, J F

    1995-01-01

    Tangier disease is a rare genetic disorder characterized by extremely low plasma levels of HDL and apo A-I, deposition of cholesteryl esters in tissues, and a high prevalence of cardiovascular disease. We examined the possibility that HDL apolipoprotein-mediated removal of cellular lipids may be defective in Tangier disease. With fibroblasts from normal subjects, purified apo A-I cleared cells of cholesteryl esters, depleted cellular free cholesterol pools available for esterification, and stimulated efflux of radiolabeled cholesterol, phosphatidylcholine, and sphingomyelin. With fibroblasts from two unrelated Tangier patients, however, apo A-I had little or no effect on any of these lipid transport processes. Intact HDL also was unable to clear cholesteryl esters from Tangier cells even though it promoted radiolabeled cholesterol efflux to levels 50-70% normal. Passive desorption of radiolabeled cholesterol or phospholipids into medium containing albumin or trypsinized HDL was normal for Tangier cells. Binding studies showed that the interaction of apo A-I with high-affinity binding sites on Tangier fibroblasts was abnormal. These results indicate that apo A-I has an impaired ability to remove cholesterol and phospholipid from Tangier fibroblasts, possibly because of a defective interaction of apo A-I with cell-surface binding sites. Failure of apo A-I to acquire cellular lipids may account for the rapid catabolism of nascent HDL particles and the low plasma HDL levels in Tangier disease. Images PMID:7615839

  14. A Model of Lipid-Free Apolipoprotein A-I Revealed by Iterative Molecular Dynamics Simulation

    PubMed Central

    Zhang, Xing; Lei, Dongsheng; Zhang, Lei; Rames, Matthew; Zhang, Shengli

    2015-01-01

    Apolipoprotein A-I (apo A-I), the major protein component of high-density lipoprotein, has been proven inversely correlated to cardiovascular risk in past decades. The lipid-free state of apo A-I is the initial stage which binds to lipids forming high-density lipoprotein. Molecular models of lipid-free apo A-I have been reported by methods like X-ray crystallography and chemical cross-linking/mass spectrometry (CCL/MS). Through structural analysis we found that those current models had limited consistency with other experimental results, such as those from hydrogen exchange with mass spectrometry. Through molecular dynamics simulations, we also found those models could not reach a stable equilibrium state. Therefore, by integrating various experimental results, we proposed a new structural model for lipid-free apo A-I, which contains a bundled four-helix N-terminal domain (1–192) that forms a variable hydrophobic groove and a mobile short hairpin C-terminal domain (193–243). This model exhibits an equilibrium state through molecular dynamics simulation and is consistent with most of the experimental results known from CCL/MS on lysine pairs, fluorescence resonance energy transfer and hydrogen exchange. This solution-state lipid-free apo A-I model may elucidate the possible conformational transitions of apo A-I binding with lipids in high-density lipoprotein formation. PMID:25793886

  15. A model of lipid-free Apolipoprotein A-I revealed by iterative molecular dynamics simulation

    DOE PAGES

    Zhang, Xing; Lei, Dongsheng; Zhang, Lei; ...

    2015-03-20

    Apolipoprotein A-I (apo A-I), the major protein component of high-density lipoprotein, has been proven inversely correlated to cardiovascular risk in past decades. The lipid-free state of apo A-I is the initial stage which binds to lipids forming high-density lipoprotein. Molecular models of lipid-free apo A-I have been reported by methods like X-ray crystallography and chemical cross-linking/mass spectrometry (CCL/MS). Through structural analysis we found that those current models had limited consistency with other experimental results, such as those from hydrogen exchange with mass spectrometry. Through molecular dynamics simulations, we also found those models could not reach a stable equilibrium state. Therefore,more » by integrating various experimental results, we proposed a new structural model for lipidfree apo A-I, which contains a bundled four-helix N-terminal domain (1–192) that forms a variable hydrophobic groove and a mobile short hairpin C-terminal domain (193–243). This model exhibits an equilibrium state through molecular dynamics simulation and is consistent with most of the experimental results known from CCL/MS on lysine pairs, fluorescence resonance energy transfer and hydrogen exchange. This solution-state lipid-free apo A-I model may elucidate the possible conformational transitions of apo A-I binding with lipids in high-density lipoprotein formation.« less

  16. A model of lipid-free Apolipoprotein A-I revealed by iterative molecular dynamics simulation

    SciTech Connect

    Zhang, Xing; Lei, Dongsheng; Zhang, Lei; Rames, Matthew; Zhang, Shengli

    2015-03-20

    Apolipoprotein A-I (apo A-I), the major protein component of high-density lipoprotein, has been proven inversely correlated to cardiovascular risk in past decades. The lipid-free state of apo A-I is the initial stage which binds to lipids forming high-density lipoprotein. Molecular models of lipid-free apo A-I have been reported by methods like X-ray crystallography and chemical cross-linking/mass spectrometry (CCL/MS). Through structural analysis we found that those current models had limited consistency with other experimental results, such as those from hydrogen exchange with mass spectrometry. Through molecular dynamics simulations, we also found those models could not reach a stable equilibrium state. Therefore, by integrating various experimental results, we proposed a new structural model for lipidfree apo A-I, which contains a bundled four-helix N-terminal domain (1–192) that forms a variable hydrophobic groove and a mobile short hairpin C-terminal domain (193–243). This model exhibits an equilibrium state through molecular dynamics simulation and is consistent with most of the experimental results known from CCL/MS on lysine pairs, fluorescence resonance energy transfer and hydrogen exchange. This solution-state lipid-free apo A-I model may elucidate the possible conformational transitions of apo A-I binding with lipids in high-density lipoprotein formation.

  17. Anti-apolipoprotein A-I antibodies and paraoxonase 1 activity in Systemic Lupus Erythematosus

    PubMed Central

    Ahmed, Mohammed Mahmoud; Elserougy, Eman Mahmoud; Al-Gazzar, Iman Ibrahim; Fikry, Iman Mohamed; Habib, Dawoud Fakhry; Younes, Khaled Mohamed; Salem, Neveen Abd El-hameed

    2013-01-01

    Systemic lupus erythematosus (SLE) patients have an increased risk of atherosclerosis. Identification of at-risk patients and the pathogenesis of atherosclerosis in SLE remain elusive. Paraoxonase 1 (PON1) and anti-apolipoprotein A-I antibody (anti-Apo A-I) appear to have a potential role in premature atherosclerosis in SLE. The aim of this work was to study PON1 activity and anti-Apo A-I antibody in SLE female patients and to demonstrate their relations to disease activity as well as disease related damage. Forty SLE female patients and 40 apparently healthy volunteers were included. Anti-Apo A-I antibodies levels and PON1 activity levels were assessed. Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) and systemic Lupus International Collaboration Clinics (SLICC)/American College of Rheumatology (ACR) damage index were preformed in all patients. Compared with controls, SLE patients showed significantly lower PON1 activity and significantly higher titers of anti-Apo A-I. Anti-Apo A-I antibody titers correlated inversely with PON1 activity. Elevated titers of anti-Apo A-I antibody and reduced PON activity were related to increased SLEDAI and (SLICC/ACR) damage index scores. We concluded that there is decreased PON1 activity and formation of anti-Apo A-I antibodies in female patients with SLE. SLE-disease activity assessed by SLEDAI and SLE disease related organ damage assessed by SLICC/ACR damage index are negatively correlated with PON1 activity and positively correlated with anti-Apo A-I antibodies. PON1 activity and anti-Apo A-I antibodies might be involved in the pathogenesis of atherosclerosis in SLE patients. PMID:26622215

  18. Contribution of polymorphisms in the apolipoprotein AI-CIII-AIV cluster to hyperlipidaemia in patients with gout.

    PubMed

    Cardona, F; Tinahones, F J; Collantes, E; Escudero, A; García-Fuentes, E; Soriguer, F J

    2005-01-01

    Studies have shown that hyperuricaemia is independently related to the insulin resistance syndrome and that polymorphisms of the apolipoprotein AI-CIII-AIV cluster are also related to insulin resistance. To study the prevalence of polymorphisms of the apolipoprotein AI-CIII-AIV cluster in persons with gout and to determine whether these polymorphisms contribute to the pathophysiology of gout or to altered lipid concentrations. Plasma cholesterol, triglycerides, uric acid, VLDL, LDL, IDL, and HDL triglycerides, cholesterol, and the renal excretion of uric acid were measured in 68 patients with gout with gout and 165 healthy subjects. Polymorphisms were studied by amplification and RFLP in all subjects, using XmnI and MspI in the apolipoprotein AI gene and SstI in the apolipoprotein CIII gene. The A allele at position -75 bp in the apolipoprotein AI gene was more common in patients with gout than in controls (p = 0.01). Levels of cholesterol, triglycerides, uric acid, basal glycaemia, and HDL cholesterol were higher in the patients (p<0.001). In the patients there was also an interaction between mutations at the two polymorphic loci studied in the apolipoprotein AI gene (p = 0.04). An absence of the mutation at position -75 bp of the apolipoprotein AI gene resulted in increased plasma triglyceride levels. Gouty patients have an altered allelic distribution in the apolipoprotein AI-CIII-AIV cluster, which could lead to changes in levels of lipoproteins. This is not caused by a single mutation but rather by a combination of different mutations.

  19. Contribution of polymorphisms in the apolipoprotein AI-CIII-AIV cluster to hyperlipidaemia in patients with gout

    PubMed Central

    Cardona, F; Tinahones, F; Collantes, E; Escudero, A; Garcia-Fuentes, E; Soriguer, F

    2005-01-01

    Background: Studies have shown that hyperuricaemia is independently related to the insulin resistance syndrome and that polymorphisms of the apolipoprotein AI-CIII-AIV cluster are also related to insulin resistance. Objective: To study the prevalence of polymorphisms of the apolipoprotein AI-CIII-AIV cluster in persons with gout and to determine whether these polymorphisms contribute to the pathophysiology of gout or to altered lipid concentrations. Methods: Plasma cholesterol, triglycerides, uric acid, VLDL, LDL, IDL, and HDL triglycerides, cholesterol, and the renal excretion of uric acid were measured in 68 patients with gout with gout and 165 healthy subjects. Polymorphisms were studied by amplification and RFLP in all subjects, using XmnI and MspI in the apolipoprotein AI gene and SstI in the apolipoprotein CIII gene. Results: The A allele at position –75 bp in the apolipoprotein AI gene was more common in patients with gout than in controls (p = 0.01). Levels of cholesterol, triglycerides, uric acid, basal glycaemia, and HDL cholesterol were higher in the patients (p<0.001). In the patients there was also an interaction between mutations at the two polymorphic loci studied in the apolipoprotein AI gene (p = 0.04). An absence of the mutation at position –75 bp of the apolipoprotein AI gene resulted in increased plasma triglyceride levels. Conclusions: Gouty patients have an altered allelic distribution in the apolipoprotein AI-CIII-AIV cluster, which could lead to changes in levels of lipoproteins. This is not caused by a single mutation but rather by a combination of different mutations. PMID:15115711

  20. Solution structure of discoidal high-density lipoprotein particles with a shortened apolipoprotein A-I.

    PubMed

    Bibow, Stefan; Polyhach, Yevhen; Eichmann, Cédric; Chi, Celestine N; Kowal, Julia; Albiez, Stefan; McLeod, Robert A; Stahlberg, Henning; Jeschke, Gunnar; Güntert, Peter; Riek, Roland

    2017-02-01

    High-density lipoprotein (HDL) particles are cholesterol and lipid transport containers. Mature HDL particles destined for the liver develop through the formation of intermediate discoidal HDL particles, which are the primary acceptors for cholesterol. Here we present the three-dimensional structure of reconstituted discoidal HDL (rdHDL) particles, using a shortened construct of human apolipoprotein A-I, determined from a combination of nuclear magnetic resonance (NMR), electron paramagnetic resonance (EPR) and transmission electron microscopy (TEM) data. The rdHDL particles feature a protein double belt surrounding a lipid bilayer patch in an antiparallel fashion. The integrity of this structure is maintained by up to 28 salt bridges and a zipper-like pattern of cation-π interactions between helices 4 and 6. To accommodate a hydrophobic interior, a gross 'right-to-right' rotation of the helices after lipidation is necessary. The structure reflects the complexity required for a shuttling container to hold a fluid lipid or cholesterol interior at a protein:lipid ratio of 1:50.

  1. Bioenergetic programming of macrophages by the apolipoprotein A-I mimetic peptide 4F.

    PubMed

    Datta, Geeta; Kramer, Philip A; Johnson, Michelle S; Sawada, Hirotaka; Smythies, Lesley E; Crossman, David K; Chacko, Balu; Ballinger, Scott W; Westbrook, David G; Mayakonda, Palgunachari; Anantharamaiah, G M; Darley-Usmar, Victor M; White, C Roger

    2015-05-01

    The apoA-I (apolipoprotein A-I) mimetic peptide 4F favours the differentiation of human monocytes to an alternatively activated M2 phenotype. The goal of the present study was to test whether the 4F-mediated differentiation of MDMs (monocyte-derived macrophages) requires the induction of an oxidative metabolic programme. 4F treatment induced several genes in MDMs that play an important role in lipid metabolism, including PPARγ (peroxisome-proliferator-activated receptor γ) and CD36. Addition of 4F was associated with a significant increase in FA (fatty acid) uptake and oxidation compared with vehicle treatment. Mitochondrial respiration was assessed by measurement of the OCR (oxygen-consumption rate). 4F increased basal and ATP-linked OCR as well as maximal uncoupled mitochondrial respiration. These changes were associated with a significant increase in ΔΨm (mitochondrial membrane potential). The increase in metabolic activity in 4F-treated MDMs was attenuated by etomoxir, an inhibitor of mitochondrial FA uptake. Finally, addition of the PPARγ antagonist T0070907 to 4F-treated MDMs reduced the expression of CD163 and CD36, cell-surface markers for M2 macrophages, and reduced basal and ATP-linked OCR. These results support our hypothesis that the 4F-mediated differentiation of MDMs to an anti-inflammatory phenotype is due, in part, to an increase in FA uptake and mitochondrial oxidative metabolism.

  2. Response to a urate-lowering diet according to polymorphisms in the apolipoprotein AI-CIII-AIV cluster.

    PubMed

    Cardona, Fernando; Tinahones, Francisco J; Collantes, Eduardo; Garcia-Fuentes, Eduardo; Escudero, Alejandro; Soriguer, Federico

    2005-05-01

    The apolipoprotein AI-CIII-AIV cluster has been associated with the response to a urate-lowering diet, and polymorphisms in the apolipoprotein CIII gene have been associated with hyperuricemia and hypertriglyceridemia. We assessed the influence of polymorphisms in the apolipoprotein AI-CIII-AIV cluster on the response to a urate-lowering diet in patients with hyperuricemia. A urate-lowering diet was followed for 2 weeks by 64 men with hyperuricemia. Plasma concentrations of triglycerides, cholesterol, glucose, and uric acid, and the uric acid clearance and 24-hour uric acid urinary excretory fraction were measured before and after the diet. The data were analyzed in association with the polymorphisms of the apolipoprotein AI-CIII-AIV gene cluster. After the urate-lowering diet, the plasma levels of triglycerides, cholesterol, glucose, and uric acid and 24-hour uric acid excretion all fell significantly. Paired sample ANOVA showed that the decrease was mainly due to the diet, except for the plasma triglycerides, which were influenced by allele X2 of the XmnI polymorphism of the apolipoprotein AI gene. The response of the biological variables to a urate-lowering diet was mainly influenced by diet. Changes in triglycerides were also influenced by the apolipoprotein AI XmnI polymorphism (p = 0.04), suggesting a gene-diet interaction (p = 0.03).

  3. Concentration and pattern changes of porcine serum apolipoprotein A-I in four different infectious diseases.

    PubMed

    Marco-Ramell, Anna; Hummel, Karin; Razzazi-Fazeli, Ebrahim; Bassols, Anna; Miller, Ingrid

    2015-02-01

    Apolipoprotein A-I (Apo A-I) is a major protein in lipid/lipoprotein metabolism and decreased serum levels have been observed in many species in response to inflammatory and infectious challenges. Little is known about the porcine homologue, therefore in this work we have characterized it through biochemical and proteomic techniques. In 2DE, porcine serum Apo A-I is found as three spots, the two more acidic ones corresponding to the mature protein, the more basic spot to the protein precursor. Despite high sequence coverage in LC-MS/MS, we did not find a sequence or PTM difference between the two mature protein species. Besides this biochemical characterization, we measured overall levels and relative species abundance of serum Apo A-I in four different viral and bacterial porcine infectious diseases. Lower overall amounts of Apo A-I were observed in Salmonella typhimurium and Escherichia coli infections. In the 2DE protein pattern, an increase of the protein precursor together with a lower level of mature protein species were detected in the porcine circovirus type 2-systemic disease and S. typhimurium infection. These results reveal that both the porcine serum Apo A-I concentration and the species pattern are influenced by the nature of the infectious disease. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Apolipoprotein B and A-I ratio predicts severe acute pancreatitis.

    PubMed

    Huh, Ji Hye; Jung, Saehyun; Cho, Seung Kook; Lee, Kyong Joo; Kim, Jae Woo

    2017-07-05

    Severe acute pancreatitis (SAP) has considerable mortality and morbidity rates. Although many indices have been developed to classify the severity of acute pancreatitis (AP), an optimal method for predicting SAP has not been identified. The ratio of apolipoprotein B to A-I (apoB/A-I) is associated with metabolic syndrome and inflammatory status. This study investigated the association between severity of AP and serum apoB/A-I ratio. Patients with AP were prospectively enrolled at Yonsei University Wonju College of Medicine from March 2015 to August 2016. The severity of AP was assessed according to the revised Atlanta classification criteria (Atlanta 2012). Of 191 patients with AP, 134 (70.2%) had mild AP, 42 (22%) had moderately severe AP, and 15 (7.9%) had SAP; apoB/A-I ratio was highest in patients with SAP (P = 0.001). The apoB/A-I ratio was positively correlated with Atlanta classification, computed tomography severity index, and Bedside index for severity of AP. The apoB/A-I ratio showed the highest predictive value for SAP in patients with AP compared with apolipoprotein B or apolipoprotein A-I alone. Serum apoB/A-I ratio appears to have value for predicting SAP in patients with AP. This article is protected by copyright. All rights reserved.

  5. In situ AFM imaging of apolipoprotein A-I directly derived from plasma HDL.

    PubMed

    Gan, Chaoye; Wang, Zhexuan; Chen, Yong

    2017-04-01

    The major apolipoproteins of plasma lipoproteins play vital roles in the structural integrity and physiological functions of lipoproteins. More than ten structural models of apolipoprotein A-I (apoA-I), the major apolipoprotein of high-density lipoprotein (HDL), have been developed successively. In these models, apoA-I was supposed to organize in a ring-shaped form. To date, however, there is no direct evidence under physiological condition. Here, atomic force microscopy (AFM) was used to in situ visualize the organization of apoA-I, which was exposed via depletion of the lipid component of plasma HDL pre-immobilized on functionalized mica sheets. For the first time, the ring-shaped coarse structure and three detailed structures (crescent-shaped, gapped "O"-shaped, and parentheses-shaped structures, respectively) of apoA-I in plasma HDL, which have the ability of binding scavenger receptors, were directly observed and quantitatively measured by AFM. The three detailed structures probably represent the different extents to which the lipid component of HDL was depleted. Data on lipid depletion of HDL may provide clues to understand lipid insertion of HDL. These data provide important information for the understanding of the structure/maturation of plasma HDL. Moreover, they suggest a powerful method for directly visualizing the major apolipoproteins of plasma lipoproteins or the protein component of lipoprotein-like lipid-protein complexes. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Apolipoprotein A-I Modulates Atherosclerosis Through Lymphatic Vessel-Dependent Mechanisms in Mice.

    PubMed

    Milasan, Andreea; Jean, Gabriel; Dallaire, François; Tardif, Jean-Claude; Merhi, Yahye; Sorci-Thomas, Mary; Martel, Catherine

    2017-09-22

    Subcutaneously injected lipid-free apoA-I (apolipoprotein A-I) reduces accumulation of lipid and immune cells within the aortic root of hypercholesterolemic mice without increasing high-density lipoprotein-cholesterol concentrations. Lymphatic vessels are now recognized as prerequisite players in the modulation of cholesterol removal from the artery wall in experimental conditions of plaque regression, and particular attention has been brought to the role of the collecting lymphatic vessels in early atherosclerosis-related lymphatic dysfunction. In the present study, we address whether and how preservation of collecting lymphatic function contributes to the protective effect of apoA-I. Atherosclerotic Ldlr(-/-) mice treated with low-dose lipid-free apoA-I showed enhanced lymphatic transport and abrogated collecting lymphatic vessel permeability in atherosclerotic Ldlr(-/-) mice when compared with albumin-control mice. Treatment of human lymphatic endothelial cells with apoA-I increased the adhesion of human platelets on lymphatic endothelial cells, in a bridge-like manner, a mechanism that could strengthen endothelial cell-cell junctions and limit atherosclerosis-associated collecting lymphatic vessel dysfunction. Experiments performed with blood platelets isolated from apoA-I-treated Ldlr(-/-) mice revealed that apoA-I decreased ex vivo platelet aggregation. This suggests that in vivo apoA-I treatment limits platelet thrombotic potential in blood while maintaining the platelet activity needed to sustain adequate lymphatic function. Altogether, we bring forward a new pleiotropic role for apoA-I in lymphatic function and unveil new potential therapeutic targets for the prevention and treatment of atherosclerosis. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

  7. Apolipoprotein A-I inhibits experimental colitis and colitis-propelled carcinogenesis.

    PubMed

    Gkouskou, K K; Ioannou, M; Pavlopoulos, G A; Georgila, K; Siganou, A; Nikolaidis, G; Kanellis, D C; Moore, S; Papadakis, K A; Kardassis, D; Iliopoulos, I; McDyer, F A; Drakos, E; Eliopoulos, A G

    2016-05-12

    In both humans with long-standing ulcerative colitis and mouse models of colitis-associated carcinogenesis (CAC), tumors develop predominantly in the distal part of the large intestine but the biological basis of this intriguing pathology remains unknown. Herein we report intrinsic differences in gene expression between proximal and distal colon in the mouse, which are augmented during dextran sodium sulfate (DSS)/azoxymethane (AOM)-induced CAC. Functional enrichment of differentially expressed genes identified discrete biological pathways operating in proximal vs distal intestine and revealed a cluster of genes involved in lipid metabolism to be associated with the disease-resistant proximal colon. Guided by this finding, we have further interrogated the expression and function of one of these genes, apolipoprotein A-I (ApoA-I), a major component of high-density lipoprotein. We show that ApoA-I is expressed at higher levels in the proximal compared with the distal part of the colon and its ablation in mice results in exaggerated DSS-induced colitis and disruption of epithelial architecture in larger areas of the large intestine. Conversely, treatment with an ApoA-I mimetic peptide ameliorated the phenotypic, histopathological and inflammatory manifestations of the disease. Genetic interference with ApoA-I levels in vivo impacted on the number, size and distribution of AOM/DSS-induced colon tumors. Mechanistically, ApoA-I was found to modulate signal transducer and activator of transcription 3 (STAT3) and nuclear factor-κB activation in response to the bacterial product lipopolysaccharide with concomitant impairment in the production of the pathogenic cytokine interleukin-6. Collectively, these data demonstrate a novel protective role for ApoA-I in colitis and CAC and unravel an unprecedented link between lipid metabolic processes and intestinal pathologies.

  8. Overexpression of apolipoprotein A-I alleviates endoplasmic reticulum stress in hepatocytes.

    PubMed

    Guo, Qing; Zhang, Can; Wang, Yutong

    2017-06-02

    Abnormal lipid metabolism may contribute to an increase in endoplasmic reticulum (ER) stress, resulting in the pathogenesis of non-alcoholic steatohepatitis. Apolipoprotein A-I (apoA-I) accepts cellular free cholesterol and phospholipids transported by ATP-binding cassette transporter A1 to generate nascent high density lipoprotein particles. Previous studies have revealed that the overexpression of apoA-I alleviated hepatic lipid levels by modifying lipid transport. Here, we examined the effects of apoA-I overexpression on ER stress and genes involved in lipogenesis in both HepG2 cells and mouse hepatocytes. Human apoA-I was overexpressed in HepG2 hepatocytes, which were then treated with 2 μg/mL tunicamycin or 500 μM palmitic acid. Eight-week-old male apoA-I transgenic or C57BL/6 wild-type mice were intraperitoneally injected with 1 mg/kg body weight tunicamycin or with saline. At 48 h after injecting, blood and liver samples were collected. The overexpression of apoA-I in the models above resulted in decreased protein levels of ER stress makers and lipogenic gene products, including sterol regulatory element binding protein 1, fatty acid synthase, and acetyl-CoA carboxylase 1. In addition, the cellular levels of triglycerides and free cholesterol also decreased. Some of gene products which are related to ER stress-associated apoptosis were also affected by apoA-I overexpression. These results suggested that apoA-I overexpression could reduce steatosis by decreasing lipid levels and by suppressing ER stress and lipogenesis in hepatocytes. ApoA-I expression could significantly reduce hepatic ER stress and lipogenesis in hepatocytes.

  9. Proteolysis of Apolipoprotein A-I by Secretory Phospholipase A2

    PubMed Central

    Cavigiolio, Giorgio; Jayaraman, Shobini

    2014-01-01

    In the acute phase of the inflammatory response, secretory phospholipase A2 (sPLA2) reaches its maximum levels in plasma, where it is mostly associated with high density lipoproteins (HDL). Overexpression of human sPLA2 in transgenic mice reduces both HDL cholesterol and apolipoprotein A-I (apoA-I) plasma levels through increased HDL catabolism by an unknown mechanism. To identify unknown PLA2-mediated activities on the molecular components of HDL, we characterized the protein and lipid products of the PLA2 reaction with HDL. Consistent with previous studies, hydrolysis of HDL phospholipids by PLA2 reduced the particle size without changing its protein composition. However, when HDL was destabilized in the presence of PLA2 by the action of cholesteryl ester transfer protein or by guanidine hydrochloride treatment, a fraction of apoA-I, but no other proteins, dissociated from the particle and was rapidly cleaved. Incubation of PLA2 with lipid-free apoA-I produced similar protein fragments in the range of 6–15 kDa, suggesting specific and direct reaction of PLA2 with apoA-I. Mass spectrometry analysis of isolated proteolytic fragments indicated at least two major cleavage sites at the C-terminal and the central domain of apoA-I. ApoA-I proteolysis by PLA2 was Ca2+-independent, implicating a different mechanism from the Ca2+-dependent PLA2-mediated phospholipid hydrolysis. Inhibition of proteolysis by benzamidine suggests that the proteolytic and lipolytic activities of PLA2 proceed through different mechanisms. Our study identifies a previously unknown proteolytic activity of PLA2 that is specific to apoA-I and may contribute to the enhanced catabolism of apoA-I in inflammation and atherosclerosis. PMID:24523407

  10. Induction of the apolipoprotein AI gene by fasting: a relationship with ketosis but not with ketone bodies.

    PubMed

    Haas, M J; Reinacher, D; Pun, K; Wong, N C; Mooradian, A D

    2000-12-01

    Apolipoprotein AI (apoAI) expression is inversely related to the incidence of atherosclerosis. ApoAI expression is also influenced by the nutritional state and diabetes. We used both cell culture and animal models to examine the effect of fasting and ketoacidosis on apoAI gene expression. Two days of food deprivation in rats increased hepatic and intestinal apoAI mRNA by 2.6- and 2.3-fold, respectively (P < .05). The absolute concentration of plasma apoAI did not change. However, the plasma apoAI concentration relative to the plasma concentration of serum proteins was increased 23% (P < .05). In fasting rats, there was a significant positive correlation between the serum beta-hydroxybutyrate concentration and hepatic or intestinal apoAI mRNA level. Despite this correlation, changes in apoAI mRNA are probably not mediated by ketone bodies, since neither hepatic nor intestinal apoAI mRNA levels were altered in rats maintained on a ketogenic diet for 10 days or treated with isobutyramide, an orally active ketone analog. In addition, the activity of the rat apoAI promoter was not altered in Hep G2 cells treated with isobutyramide or fatty acids or exposed to hypoglycemic conditions, while dexamethasone increased promoter activity 1.9-fold (P < .05). These data indicate that metabolic changes other than ketone bodies, such as an increase in plasma glucocorticoids, may account for starvation-induced expression of apoAI.

  11. Local Vascular Gene Therapy With Apolipoprotein A-I to Promote Regression of Atherosclerosis.

    PubMed

    Wacker, Bradley K; Dronadula, Nagadhara; Zhang, Jingwan; Dichek, David A

    2017-02-01

    Gene therapy, delivered directly to the blood vessel wall, could potentially prevent atherosclerotic lesion growth and promote atherosclerosis regression. Previously, we reported that a helper-dependent adenoviral (HDAd) vector expressing apolipoprotein A-I (apoA-I) in carotid endothelium of fat-fed rabbits reduced early (4 weeks) atherosclerotic lesion growth. Here, we tested whether the same HDAd-delivered to the existing carotid atherosclerotic lesions-could promote regression. Rabbits (n=26) were fed a high-fat diet for 7 months, then treated with bilateral carotid gene transfer. One carotid was infused with an HDAd expressing apoA-I (HDAdApoAI) and the other with a control nonexpressing HDAd (HDAdNull). The side with HDAdApoAI was randomized. Rabbits were then switched to regular chow, lowering their plasma cholesterols by over 70%. ApoA-I mRNA and protein were detected in HDAdApoAI-transduced arteries. After 7 weeks of gene therapy, compared with HDAdNull-treated arteries in the same rabbits, HDAdApoAI-treated arteries had significantly less vascular cell adhesion molecule-1 expression (28%; P=0.04) along with modest but statistically insignificant trends toward decreased intimal lesion volume, lipid and macrophage content, and intercellular adhesion molecule-1 expression (9%-21%; P=0.1-0.4). Post hoc subgroup analysis of rabbits with small-to-moderate-sized lesions (n=20) showed that HDAdApoAI caused large reductions in lesion volume, lipid content, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 expression (30%-50%; P≤0.04 for all). Macrophage content was reduced by 30% (P=0.06). There was a significant interaction (P=0.02) between lesion size and treatment efficacy. Even when administered on a background of aggressive lowering of plasma cholesterol, local HDAdApoAI vascular gene therapy may promote rapid regression of small-to-moderate-sized atherosclerotic lesions. © 2016 American Heart Association, Inc.

  12. Effect of an isoenergetic traditional Mediterranean diet on apolipoprotein A-I kinetic in men with metabolic syndrome

    USDA-ARS?s Scientific Manuscript database

    The impact of the Mediterranean diet (MedDiet) on high-density lipoprotein (HDL) kinetics has not been studied to date. The objective of this study was therefore to investigate the effect of the MedDiet in the absence of changes in body weight on apolipoprotein (apo) A-I kinetic in men with metaboli...

  13. Apolipoprotein A-I and A-I mimetic peptides: a role in atherosclerosis

    PubMed Central

    Getz, Godfrey S; Reardon, Catherine A

    2011-01-01

    Cardiovascular disease remains a major cause of morbidity and mortality in the westernized world. Atherosclerosis is the underlying cause of most cardiovascular diseases. Atherosclerosis is a slowly evolving chronic inflammatory disorder involving the intima of large and medium sized arteries that is initiated in response to high plasma lipid levels, especially LDL. Cells of both the innate and adaptive immunity are involved in this chronic inflammation. Although high plasma LDL levels are a major contributor to most stages of the evolution of atherosclerosis, HDL and its major protein apoA-I possess properties that attenuate and may even reverse atherosclerosis. Two major functions are the ability to induce the efflux of cholesterol from cells, particularly lipid-loaded macrophages, in the artery wall for transfer to the liver, a process referred to as reverse cholesterol transport, and the ability to attenuate the pro-inflammatory properties of LDL. The removal of cellular cholesterol from lipid-loaded macrophages may also be anti-inflammatory. One of the most promising therapies to enhance the anti-atherogenic, anti-inflammatory properties of HDL is apoA-I mimetic peptides. Several of these peptides have been shown to promote cellular cholesterol efflux, attenuate the production of pro-inflammatory cytokines by macrophages, and to attenuate the pro-inflammatory properties of LDL. This latter effect may be related to their high affinity for oxidized lipids present in LDL. This review discusses the functional properties of the peptides and their effect on experimental atherosclerosis and the results of initial clinical studies in humans. PMID:22096372

  14. Endotoxin contamination of apolipoprotein A-I: effect on macrophage proliferation--a cautionary tale.

    PubMed

    Jin, Xueting; Xu, Qing; Champion, Keith; Kruth, Howard S

    2015-05-01

    This technical report addresses the problem of endotoxin contamination of apolipoprotein reagents. Using a bromodeoxyuridine incorporation cell proliferation assay, we observed that human plasma ApoA-I as low as 1 μg/ml resulted in a >90% inhibition in macrophage proliferation. However, not all ApoA-I from different sources showed this effect. We considered the possibility that endotoxin contamination of the apolipoproteins contributed to the differential inhibition of macrophage cell proliferation. Endotoxin alone very potently inhibited macrophage proliferation (0.1 ng/ml inhibited macrophage proliferation>90%). Measurement of endotoxin levels in the apolipoprotein products, including an analysis of free versus total endotoxin, the latter which included endotoxin that was masked due to binding to protein, suggested that free endotoxin mediated inhibition of macrophage proliferation. Despite the use of an advanced endotoxin removal procedure and agents commonly used to inhibit endotoxin action, the potency of endotoxin precluded successful elimination of endotoxin effect. Our findings show that endotoxin contamination can significantly influence apparent apolipoprotein-mediated cell effects (or effects of any other biological products), especially when these products are tested on highly endotoxin-sensitive cells, such as macrophages.

  15. Endotoxin Contamination of Apolipoprotein A-I: Effect on Macrophage Proliferation – A Cautionary Tale

    PubMed Central

    Jin, Xueting; Xu, Qing; Champion, Keith; Kruth, Howard S.

    2015-01-01

    This technical report addresses the problem of endotoxin contamination of apolipoprotein reagents. Using a bromodeoxyuridine incorporation cell proliferation assay, we observed that human plasma ApoA-I as low as 1 μg/ml resulted in a >90% inhibition in macrophage proliferation. However, not all ApoA-I from different sources showed this effect. We considered the possibility that endotoxin contamination of the apolipoproteins contributed to the differential inhibition of macrophage cell proliferation. Endotoxin alone very potently inhibited macrophage proliferation (0.1 ng/ml inhibited macrophage proliferation >90%). Measurement of endotoxin levels in the apolipoprotein products, including an analysis of free versus total endotoxin, the latter which included endotoxin that was masked due to binding to protein, suggested that free endotoxin mediated inhibition of macrophage proliferation. Despite the use of an advanced endotoxin removal procedure and agents commonly used to inhibit endotoxin action, the potency of endotoxin precluded successful elimination of endotoxin effect. Our findings show that endotoxin contamination can significantly influence apparent apolipoprotein-mediated cell effects (or effects of any other biological products), especially when these products are tested on highly endotoxin-sensitive cells, such as macrophages. PMID:25778625

  16. Inducing apolipoprotein A-I synthesis to reduce cardiovascular risk: from ASSERT to SUSTAIN and beyond.

    PubMed

    Di Bartolo, Belinda A; Scherer, Daniel J; Nicholls, Stephen J

    2016-12-01

    Increasing attention has focused on efforts to promote the biological activities of high-density lipoproteins (HDL) in order to reduce cardiovascular risk. Targeting apolipoprotein A-I (apoA-I), the major protein carried on HDL particles, represents an attractive approach to promoting HDL by virtue of its ability to increase endogenous synthesis of functional HDL particles. A number of pharmacological strategies that target apoA-I, including upregulation of its production with the bromodomain and extraterminal (BET) protein inhibitor RVX-208, development of short peptide sequences that mimic its action, and administration as a component of reconstituted HDL particles, have undergone clinical development. The impact of these approaches on cardiovascular biomarkers will be reviewed.

  17. Membrane effects of N-terminal fragment of apolipoprotein A-I: a fluorescent probe study.

    PubMed

    Trusova, Valeriya; Gorbenko, Galyna; Girych, Mykhailo; Adachi, Emi; Mizuguchi, Chiharu; Sood, Rohit; Kinnunen, Paavo; Saito, Hiroyuki

    2015-03-01

    The binding of monomeric and aggregated variants of 1-83 N-terminal fragment of apolipoprotein A-I with substitution mutations G26R, G26R/W@8, G26R/W@50 and G26R/W@72 to the model lipid membranes composed of phosphatidylcholine and its mixture with cholesterol has been investigated using fluorescent probes pyrene and Laurdan. Examination of pyrene spectral behavior did not reveal any marked influence of apoA-I mutants on the hydrocarbon region of lipid bilayer. In contrast, probing the membrane effects by Laurdan revealed decrease in the probe generalized polarization in the presence of aggregated proteins. suggesting that oligomeric and fibrillar apoA-I species induce increase in hydration degree and reduction of lipid packing density in the membrane interfacial region. These findings may shed light on molecular details of amyloid cytotoxicity.

  18. C1QBP is upregulated in colon cancer and binds to apolipoprotein A-I.

    PubMed

    Kim, Kun; Kim, Min-Jeong; Kim, Kyung-Hee; Ahn, Sun-A; Kim, Jong Heon; Cho, Jae Youl; Yeo, Seung-Gu

    2017-05-01

    The present study aimed to investigate the expression of complement component 1, q subcomponent-binding protein (C1QBP) in colon cancer cells, and identify proteins that interact with C1QBP. Total proteins were extracted from both the tumor and normal tissues of 22 patients with colon cancer and analyzed using liquid chromatography-mass spectrometry (LC-MS) to identify proteins that were differentially-expressed in tumor tissues. C1QBP overexpression was induced in 293T cells using a pFLAG-CMV2 expression vector. Overexpressed FLAG-tagged C1QBP protein was then immunoprecipitated using anti-FLAG antibodies and C1QBP-interacting proteins were screened using LC-MS analysis of the immunoprecipitates. The C1QBP-interacting proteins were confirmed using reverse-immunoprecipitation and the differential expression of C1QBP in tissues and cell lines was confirmed using western blot analysis. LC-MS analysis revealed that C1QBP exhibited a typical tumor expression pattern. Two immune-reactive signals (33 and 14 kDa) were detected in normal and tumor tissues from 19 patients. Furthermore, 14 kDa C1QBP protein was upregulated in the tumors of 15 patients. In total, 39 proteins were identified as candidate C1QBP-interacting proteins, and an interaction between C1QBP and apolipoprotein A-I was confirmed. The present study indicates that C1QBP is involved in colon cancer carcinogenesis, and that the mechanisms underlying the established anti-tumor properties of apolipoprotein A-I may include interacting with and inhibiting the activity of C1QBP.

  19. Isolation and function analysis of apolipoprotein A-I gene response to virus infection in grouper.

    PubMed

    Wei, Jingguang; Gao, Pin; Zhang, Ping; Guo, Minglan; Xu, Meng; Wei, Shina; Yan, Yang; Qin, Qiwei

    2015-04-01

    Apolipoproteins, synthesized mainly in liver and intestine and bounded to lipids, play important roles in lipid transport and uptake through the circulation system. In this study, an apolipoprotein A-I gene homologue was cloned from orange-spotted grouper Epinephelus coioides (designed as Ec-ApoA-I) by rapid amplification of cDNA ends (RACE) PCR. The full-length cDNA of Ec-ApoA-I was comprised of 1278 bp with a 792 bp open reading frame (ORF) that encodes a putative protein of 264 amino acids. Quantitative real-time PCR (qPCR) analysis revealed that Ec-ApoA-I was abundant in liver and intestine, and the expression in liver was significantly (P < 0.01) up-regulated after the stimulation of LPS, Poly(I:C), Vibrio alginolyticus, and Singapore grouper iridovirus (SGIV). Recombinant Ec-ApoA-I (rEc-ApoA-I) was produced in Escherichia coli BL21 (DE3) expression system exhibited bacteriolyticactivity against Microcococcus lysodeikticus and Aeromonas hydrophila. Intracellular localization revealed that Ec-ApoA-I distributed in both cytoplasm and nucleus, and predominantly in the cytoplasm. Overexpression of Ec-ApoA-I in grouper Brain (GB) cells could inhibit the replication of SGIV. These results together indicated that Ec-ApoA-I perhaps is involved in the responses to bacterial and viral challenge. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Serum apolipoproteins A-I and B in 2,854 children from a biracial community: Bogalusa Heart Study.

    PubMed

    Srinivasan, S R; Freedman, D S; Sharma, C; Webber, L S; Berenson, G S

    1986-08-01

    Serum apolipoprotein A-I (apo A-I) and apolipoprotein B (apo B) profiles were examined in 2,854 children, 5 to 17 years of age, from a total biracial community. Black boys had higher apo A-I levels than white boys (P less than .001), whereas girls showed no such race-related difference. Black-white difference in apo A-I persisted among boys with similar triglyceride levels provided that triglyceride levels were high. The ratio of high-density lipoprotein cholesterol (HDL-C)/apo A-I was higher in black than in white children, irrespective of sex (P less than .001). Only black children showed sex-related differences for apo A-I (boys greater than girls, P less than .05). Sex-related differences were seen in white children for HDL-C/apo A-I ratio (boys greater than girls, P less than .001) and in children of both races for apoB (girls greater than boys, P less than .01). Age-related changes were more apparent for apo A-I and HDL-C/apo A-I ratio than for apo B. A progressive decrease in apo A-I was noted during sexual maturation only in white boys. The magnitude of inverse association of apo B to HDL-C was less strong in black children (P less than .01). Although apo A-I was inversely correlated with very low-density lipoprotein cholesterol and triglycerides in white children, no association was noted in black children. These findings are indicative of intrinsic metabolic differences among the race-sex groups, resulting in variability in lipoprotein composition and levels and atherogenic potential.

  1. Serum concentrations of cholesterol, apolipoprotein A-I and apolipoprotein B in a total of 1694 meat-eaters, fish-eaters, vegetarians and vegans.

    PubMed

    Bradbury, K E; Crowe, F L; Appleby, P N; Schmidt, J A; Travis, R C; Key, T J

    2014-02-01

    The objective of this study was to describe serum lipid concentrations, including apolipoproteins A-I and B, in different diet groups. A cross-sectional analysis of a sample of 424 meat-eaters, 425 fish-eaters, 423 vegetarians and 422 vegans, matched on sex and age, from the European Prospective Investigation into Cancer and Nutrition-Oxford cohort. Serum concentrations of total, and high-density lipoprotein (HDL) cholesterol, as well as apolipoproteins A-I and B were measured, and serum non-HDL cholesterol was calculated. Vegans had the lowest body mass index (BMI) and the highest and lowest intakes of polyunsaturated and saturated fat, respectively. After adjustment for age, alcohol and physical activity, compared with meat-eaters, fish-eaters and vegetarians, serum concentrations of total and non-HDL cholesterol and apolipoprotein B were significantly lower in vegans. Serum apolipoprotein A-I concentrations did not differ between the diet groups. In males, the mean serum total cholesterol concentration was 0.87 mmol/l lower in vegans than in meat-eaters; after further adjustment for BMI this difference was 0.76 mmol/l. In females, the difference in total cholesterol between these two groups was 0.6 mmol/l, and after further adjustment for BMI was 0.55 mmol/l. [corrected]. In this study, which included a large number of vegans, serum total cholesterol and apolipoprotein B concentrations were lower in vegans compared with meat-eaters, fish-eaters and vegetarians. A small proportion of the observed differences in serum lipid concentrations was explained by differences in BMI, but a large proportion is most likely due to diet.

  2. Serum concentrations of cholesterol, apolipoprotein A-I, and apolipoprotein B in a total of 1 694 meat-eaters, fish-eaters, vegetarians, and vegans

    PubMed Central

    Bradbury, Kathryn E; Crowe, Francesca L; Appleby, Paul N; Schmidt, Julie A; Travis, Ruth C; Key, Timothy J

    2013-01-01

    BACKGROUND The objective of this study was to describe serum lipid concentrations, including apolipoproteins A-I and B, in different diet groups. METHODS A cross-sectional analysis of a sample of 424 meat-eaters, 425 fish-eaters, 423 vegetarians, and 422 vegans, matched on sex and age, from the European Prospective Investigation into Cancer and Nutrition (EPIC)-Oxford cohort. Serum concentrations of total, and HDL cholesterol, as well as apolipoproteins A-I and B were measured, and serum non-HDL cholesterol was calculated. RESULTS Vegans had the lowest BMI, and the highest and lowest intakes of polyunsaturated and saturated fat, respectively. After adjustment for age, alcohol and physical activity, compared to meat-eaters, fish-eaters and vegetarians, serum concentrations of total and non-HDL cholesterol, and apolipoprotein B were significantly lower in vegans. Serum apolipoprotein A-I concentrations did not differ between the diet groups. In males, the mean serum total cholesterol concentration was 0.87 nmol/L lower in vegans than in meat-eaters; after further adjustment for BMI this difference was 0.76 nmol/L. In females, the difference in total cholesterol between these two groups was 0.60 nmol/L, and after further adjustment for BMI was 0.55 nmol/L. CONCLUSIONS In this study, which included a large number of vegans, serum total cholesterol and apolipoprotein B concentrations were lower in vegans compared to meat-eaters, fish-eaters and vegetarians. A small proportion of the observed differences in serum lipid concentrations was explained by differences in BMI, but a large proportion is most likely due to diet. PMID:24346473

  3. Apolipoprotein AI-CIII-AIV gene cluster polymorphisms in relation to cholesterol gallstone.

    PubMed

    Yao, You Gui; Qiu, Xiong; Ma, Ming Kun; Pu, Dao Sheng; Xiao, Lu Jia

    2007-02-01

    To investigate the frequency of variants at Xmn I, Msp I sites of apolipoprotein (Apo), A I-CIII-AIV gene cluster, and its relation to cholesterol gallstones in Chinese patients. Restriction fragment length polymorphisms (RFLP) at Xmn I, Msp I sites of ApoAI-CIII-AIV gene cluster were studied using a polymerase chain reaction (PCR) in 161 patients with cholesterol gallstones and 94 healthy subjects from a Chinese population in Sichuan Province. In both the cholesterol gallstone group and the healthy control group, X1 and M1 alleles were the major alleles and homozygous X1X1 and M1M1 genotypes were the most frequent. However, the frequency of X2 allele mutation in female patients of the cholesterol gallstones group was significantly higher than that in women in the healthy control group (P<0.05), but no difference was found in the frequency of M2 alleles mutation (P>0.05). The data showed that Xmn I RFLP of ApoAI-CIII-AIV gene cluster is associated to some extent with cholesterol gallstones in female Chinese patients.

  4. RVX-208, a stimulator of apolipoprotein AI gene expression for the treatment of cardiovascular diseases.

    PubMed

    McNeill, Eileen

    2010-03-01

    RVX-208 (RVX-000222) is a first-in-class, orally active, small-molecule stimulator of apolipoprotein (APO)AI gene expression, which is being developed by Resverlogix Corp for the potential treatment of cardiovascular diseases, in particular atherosclerosis and coronary artery disease. In vitro, RVX-208 stimulated APOAI transcription and was associated with dose-dependent increases in apoAI mRNA and protein. Toxicity studies in animals and phase I/II clinical trials have indicated that RVX-208 is safe and well tolerated in multiple dosing regimens. Plasma exposure to RVX-208 was dose-dependent following single or multiple oral doses and the drug was readily absorbed. In healthy volunteers and patients with low HDL-cholesterol levels, RVX-208 increased total HDL as well as the alpha- and pre-beta HDL fractions that are important substrates in the reverse cholesterol transport pathway. The results of further phase II trials are eagerly awaited to determine whether RVX-208 can deliver plaque regression via improvements in the plasma HDL profile of patients. In addition, a phase I trial indicated that RVX-208 may have potential for the removal of beta-amyloid plaques in Alzheimer's disease and this will be further assessed in an ongoing phase I/II clinical trial.

  5. Apolipoprotein A-I and Paraoxonase-1 Are Potential Blood Biomarkers for Ischemic Stroke Diagnosis.

    PubMed

    Walsh, Kyle B; Hart, Kimberly; Roll, Susan; Sperling, Matthew; Unruh, Dusten; Davidson, W Sean; Lindsell, Christopher J; Adeoye, Opeolu

    2016-06-01

    Blood biomarkers for ischemic and hemorrhagic stroke diagnosis remain elusive. Recent investigations suggested that apolipoprotein (Apo), matrix metalloproteinase (MMP), and paraoxonase-1 may be associated with stroke. We hypothesized that Apo A-I, Apo C-I, Apo C-III, MMP-3, MMP-9, and paraoxonase-1 are differentially expressed in ischemic stroke, hemorrhagic stroke, and controls. In a single-center prospective observational study, consecutive stroke cases were enrolled if blood samples were obtainable within 12 hours of symptom onset. Age- (±5 years), race-, and sex-matched controls were recruited. Multiplex assays were used to measure protein levels. The Wilcoxon signed-rank test and the Mann-Whitney U-test were used to compare biomarker values between ischemic stroke patients and controls, hemorrhagic stroke patients and controls, and ischemic and hemorrhagic stroke patients. The 95% confidence intervals (CIs) for the difference of 2 medians were calculated. Fourteen ischemic stroke case-control pairs and 23 intracerebral hemorrhage (ICH) case-control pairs were enrolled. Median Apo A-I levels were lower in ischemic stroke cases versus controls (140 mg/dL versus 175 mg/dL, difference of 35 mg/dL, 95% CI -54 to -16) and in ischemic stroke versus ICH cases (140 mg/dL versus 180 mg/dL, difference of 40 mg/dL, 95% CI -57 to -23). Median paraoxonase-1 was lower in ischemic stroke cases than in both ICH cases and matched controls. Median Apo C-I was slightly lower in ischemic stroke cases than in ICH cases. There were no differences between groups for MMP-3, MMP-9, and Apo C-III. Apo A-I and paraoxonase-1 levels may be clinically useful for ischemic stroke diagnosis and for differentiating between ischemic and hemorrhagic strokes. Copyright © 2016 National Stroke Association. Published by Elsevier Inc. All rights reserved.

  6. [Screening and identification of apolipoprotein A-I as a potential marker for hepatoblastoma in children].

    PubMed

    Guo, Li-Hua; Zhao, Wei; Zhang, Jun-Jie; Zhang, Qian; Fan, Ying-Zhong; Wang, Jia-Xiang

    2016-12-01

    To screen and identify serum biomarkers for childhood hepatoblastoma (HB). The serum samples from 30 children with hepatoblastoma (HB), 20 children with systemic inflammatory response syndrome, and 20 normal children were treated with magnetic bead-based weak cation exchange chromatography. The platform of surface-enhanced laser desorption/ionization-time of flight-mass spectrometry (SELDI-TOF-MS) was used to eliminate the interference of inflammatory factors and to screen out the differentially expressed proteins in serum between tumor group and normal group. After the purification and separation of target proteins were performed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, matrix-assisted laser desorption/ionization-time of flight-mass spectrometry was used to determine their amino acid sequences. The SwissProt database was searched for matched proteins. Finally, real-time PCR and ELISA were used to verify and measure the expression of target proteins. After SELDI-TOF-MS was used for screening and elimination of the interference of inflammatory factors, a differentially expression protein with a mass-to-charge ratio of 9 348 Da was found in serum between HB group and normal group, and the HB group had significantly lower expression of this protein than the normal group (p<0.05). This protein was identified as apolipoprotein A-1 (Apo A-I). Real-time PCR and ELISA verified the low mRNA and protein expression of Apo A-I in serum in the HB group and high expression in serum in the normal group. Apo A-I can be used as a non-inflammatory protein marker for HB and has a certain value in the early diagnosis of HB.

  7. Heterogeneous expression of apolipoprotein-E by human macrophages

    PubMed Central

    Tedla, Nicodemus; Glaros, Elias N; Brunk, Ulf T; Jessup, Wendy; Garner, Brett

    2004-01-01

    Apolipoprotein-E (apoE) is expressed at high levels by macrophages. In addition to its role in lipid transport, macrophage-derived apoE plays an important role in immunoregulation. Previous studies have identified macrophage subpopulations that differ substantially in their ability to synthesize specific cytokines and enzymes, however, potential heterogeneous macrophage apoE expression has not been studied. Here we examined apoE expression in human THP-1 macrophages and monocyte-derived macrophages (MDM). Using immunocytochemistry and flow cytometry methods we reveal a striking heterogeneity in macrophage apoE expression in both cell types. In phorbol-ester-differentiated THP-1 macrophages, 5% of the cells over-expressed apoE at levels more than 50-fold higher than the rest of the population. ApoE over-expressing THP-1 macrophages contained condensed/fragmented nuclei and increased levels of activated caspase-3 indicating induction of apoptosis. In MDM, 3–5% of the cells also highly over-expressed apoE, up to 50-fold higher than the rest of the population; however, this was not associated with obvious nuclear alterations. The apoE over-expressing MDM were larger, more granular, and more autofluorescent than the majority of cells and they contained numerous vesicle-like structures that appeared to be coated by apoE. Flow cytometry experiments indicated that the apoE over-expressing subpopulation of MDM were positive for CD14, CD11b/Mac-1 and CD68. These observations suggest that specific macrophage subpopulations may be important for apoE-mediated immunoregulation and clearly indicate that subpopulation heterogeneity should be taken into account when investigating macrophage apoE expression. PMID:15500620

  8. Effect of body mass index on apolipoprotein A-I kinetics in middle-aged men and postmenopausal women.

    PubMed

    Welty, Francine K; Lichtenstein, Alice H; Lamon-Fava, Stefania; Schaefer, Ernst J; Marsh, Julian B

    2007-07-01

    The effect of body mass index (BMI) and obesity on apolipoprotein (apo) A-I levels and kinetics was examined by gender. Apo A-I kinetics were determined with a primed, constant infusion of deuterated leucine in the fed state in 19 men and 13 postmenopausal women. Compared with nonobese men, nonobese women had a higher level of high-density lipoprotein cholesterol (HDL-C) and apo A-I due to a 48% higher apo A-I production rate (PR) (P = .05). Obesity had no significant effects on apo A-I kinetics in women. In contrast, compared with nonobese men, obese men had a 9% lower apo A-I level due to a 64% higher fractional catabolic rate (FCR) partially offset by a 47% higher PR. Obese women had a 52% higher HDL-C than obese men (50 vs 33 mg/dL, respectively; P = .012), a finding related to the faster apo A-I FCR in obese men. BMI was directly correlated with apo A-I FCR (r = 0.84, P < .001) and PR (r = 0.79, P < .001) in men but not in women. Sixty-two percent of the variability in PR and 71% of the variability in FCR were due to BMI in men and only 3% and 23%, respectively, in women. In conclusion, BMI has a significant effect on apo A-I PR and FCR in men but not in women.

  9. Interactions between lipid-free apolipoprotein-AI and a lipopeptide incorporating the RGDS cell adhesion motif

    NASA Astrophysics Data System (ADS)

    Castelletto, V.; Hamley, I. W.; Reza, M.; Ruokolainen, J.

    2014-11-01

    The interaction of a designed bioactive lipopeptide C16-GGGRGDS, comprising a hexadecyl lipid chain attached to a functional heptapeptide, with the lipid-free apoliprotein, Apo-AI, is examined. This apolipoprotein is a major component of high density lipoprotein and it is involved in lipid metabolism and may serve as a biomarker for cardiovascular disease and Alzheimers' disease. We find via isothermal titration calorimetry that binding between the lipopeptide and Apo-AI occurs up to a saturation condition, just above equimolar for a 10.7 μM concentration of Apo-AI. A similar value is obtained from circular dichroism spectroscopy, which probes the reduction in α-helical secondary structure of Apo-AI upon addition of C16-GGGRGDS. Electron microscopy images show a persistence of fibrillar structures due to self-assembly of C16-GGGRGDS in mixtures with Apo-AI above the saturation binding condition. A small fraction of spheroidal or possibly ``nanodisc'' structures was observed. Small-angle X-ray scattering (SAXS) data for Apo-AI can be fitted using a published crystal structure of the Apo-AI dimer. The SAXS data for the lipopeptide/Apo-AI mixtures above the saturation binding conditions can be fitted to the contribution from fibrillar structures coexisting with flat discs corresponding to Apo-AI/lipopeptide aggregates.

  10. Expression and purification of recombinant apolipoprotein A-I Zaragoza (L144R) and formation of reconstituted HDL particles.

    PubMed

    Fiddyment, Sarah; Barceló-Batllori, Sílvia; Pocoví, Miguel; García-Otín, Angel-Luis

    2011-11-01

    Apolipoprotein A-I Zaragoza (L144R) (apo A-I Z), has been associated with severe hypoalphalipoproteinemia and an enhanced effect of high density lipoprotein (HDL) reverse cholesterol transport. In order to perform further studies with this protein we have optimized an expression and purification method of recombinant wild-type apo A-I and apo A-I Z and produced mimetic HDL particles with each protein. An pET-45 expression system was used to produce N-terminal His-tagged apo A-I, wild-type or mutant, in Escherichia coli BL21 (DE3) which was subsequently purified by affinity chromatography in non-denaturing conditions. HDL particles were generated via a modified sodium cholate method. Expression and purification of both proteins was verified by SDS-PAGE, MALDI-TOF MS and immunochemical procedures. Yield was 30mg of purified protein (94% purity) per liter of culture. The reconstituted HDL particles checked via non-denaturing PAGE showed high homogeneity in their size when reconstituted both with wild-type apo A-I and apo A-I Z. An optimized system for the expression and purification of wild-type apo A-I and apo A-I Z with high yield and purity grade has been achieved, in addition to their use in reconstituted HDL particles, as a basis for further studies.

  11. Correlations Between Serum Apolipoprotein A-I and Formation of Vocal Cord Polyp.

    PubMed

    Zhang, Hui-Ping; Zhang, Rong

    2017-05-01

    This study aims to investigate the correlations between serum apolipoprotein A-I (ApoA-I) and the formation of vocal cord polyps (VCPs). This study used the nonmatched case-control study method. The serum total cholesterol (TC), triglyceride, high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol, ApoA-I, apolipoprotein B (ApoB), and ApoA-I/ApoB levels of 89 VCP patients and 87 normal volunteers were compared. Additionally, such VCP-related factors as excessive vocal use, vocal abuse, smoking, drinking, and the size of VCPs were analyzed. The two groups did not significantly differ with regard to triglyceride, low-density lipoprotein cholesterol, ApoB, and ApoA-I/ApoB levels (P > 0.05), whereas they did significantly differ with regard to TC, HDL-C, and ApoA-I levels (P < 0.05) according to independent t tests. Logistic regression analysis showed that excessive vocal use and vocal abuse were risk factors for VCPs (P < 0.05), with odds ratio values of 5.675 and 12.781, respectively. The ApoA-I level was negatively associated with VCPs (P < 0.05), with an odds ratio of 0.511; however, TC and HDL-C were not associated with the formation of VCPs (P > 0.05). The size of VCPs in females was negatively correlated with the serum ApoA-I level (r = -0.349, P = 0.032), whereas that in males was not (P > 0.05). As the serum ApoA-I level was negatively correlated with the formation of VCPs, ApoA-I may reduce the risk of VCPs. These findings may facilitate the prevention and treatment of VCPs. Copyright © 2017. Published by Elsevier Inc.

  12. Reduced Cerebrospinal Fluid Concentration of Apolipoprotein A-I in Patients with Alzheimer's Disease.

    PubMed

    Johansson, Per; Almqvist, Erik G; Bjerke, Maria; Wallin, Anders; Johansson, Jan-Ove; Andreasson, Ulf; Blennow, Kaj; Zetterberg, Henrik; Svensson, Johan

    2017-01-01

    Apolipoprotein E (ApoE) has been extensively studied in Alzheimer's disease (AD), but little is known of apolipoprotein A-I (ApoA-I) in cerebrospinal fluid (CSF). Plasma lipids as well as ApoA-I and ApoE in plasma and CSF were determined and related to Mini-Mental State Examination (MMSE) score, APOE genotype, and CSF AD biomarkers. Consecutive patients with AD (n = 29), stable mild cognitive impairment (n = 13), other dementias (n = 14), and healthy controls (n = 18) were included at a single center. AD patients had higher plasma triglycerides and lower CSF ApoA-I concentration than controls (both p < 0.05). CSF ApoE concentration was reduced in other dementias (p < 0.01). In AD as well as other dementias, the ratios between CSF and plasma concentrations of both ApoA-I and ApoE were lower than those in the controls. ApoA-I and ApoE in plasma and CSF were not influenced by APOEɛ4 allele distribution. In the total study population (n = 74), CSF ApoA-I correlated positively with MMSE score (r = 0.26, p < 0.05) and negatively with CSF P-tau (r = -0.25, p < 0.05). CSF ApoE correlated positively with CSF concentrations of T-tau and P-tau in the total study population and in AD patients. CSF ApoA-I was reduced in AD patients and associated with measures of cognitive function and AD disease status. The mechanisms underlying the decreased CSF:plasma ratios of ApoA-I and ApoE in AD and other dementias need to be explored in further studies.

  13. Apolipoprotein A-I and platelet factor 4 are biomarkers for infliximab response in rheumatoid arthritis

    PubMed Central

    Trocmé, Candice; Marotte, Hubert; Baillet, Athan; Pallot-Prades, Béatrice; Garin, Jérome; Grange, Laurent; Miossec, Pierre; Tebib, Jacques; Berger, François; Nissen, Michael J.; Juvin, Robert; Morel, Françoise; Gaudin, Philippe

    2009-01-01

    Objectives The use of biologics such as infliximab has dramatically improved the treatment of rheumatoid arthritis (RA). However, factors predictive of therapeutic response need to be identified. A proteomic study was performed prior to infliximab therapy to identify a panel of candidate protein biomarkers of RA predictive of treatment response. Methods Plasma profiles of 60 RA patients (28 non-responders ACR 20 negative and 32 responders ACR 70 positive to infliximab) were studied by SELDI-TOF-MS technology on two types of arrays, an anion exchange array (SAX2) and a nickel affinity array (IMAC3-Ni). Biomarker characterization was carried out using classical biochemical methods (purification by ammonium sulfate precipitation or metal affinity chromatography) and identification by MALDI-TOF analysis. Results Two distinct protein profiles were observed on both arrays and several proteins were differentially expressed in both patient populations. Five proteins at 3.86, 7.77, 7.97, 8.14 and 74.07 kDa were overexpressed in the non-responder group, whereas one at 28 kDa was increased in the responder population (sensitivity > 56%, specificity > 77.5%). Moreover combination of several biomarkers improved both the sensitivity and specificity of the detection of patient response to over 97%. The 28 kDa protein was characterized as apolipoprotein A-I and the 7.77 kDa biomarker was identified as platelet factor 4. Conclusions We characterized six plasma biomarkers, enabling the detection of patient response to infliximab with high sensitivity and specificity. Apolipoprotein A-1 was predictive of a good response to infliximab, whereas platelet factor 4 was associated with non-responders. PMID:18664547

  14. Mosaic Interdigitated Structure in Nanoparticle-Templated Phospholipid Bilayer Supports Partial Lipidation of Apolipoprotein A-I.

    PubMed

    Sun, Wangqiang; Wu, Weiqiang; McMahon, Kaylin M; Rink, Jonathan S; Thaxton, C Shad

    2016-06-01

    Using gold nanoparticle-templated high-density lipoprotein-like particles as a model, the nanoparticle-templated phospholipid bilayer is studied from the bottom-up. Data support the phospholipids have a mosaic interdigitated structure. The discontinuous lipid milieu supports partial lipidation of apolipoprotein A-I, different from an ordinary phospholipid bilayer, suggesting that synergy between nanoparticle templates and bound phospholipid layers can modulate amphiphilic proteins for desired functions.

  15. The Concentration of Apolipoprotein A-I Decreases during Experimentally Induced Acute-Phase Processes in Pigs

    PubMed Central

    Carpintero, R.; Piñeiro, M.; Andrés, M.; Iturralde, M.; Alava, M. A.; Heegaard, P. M. H.; Jobert, J. L.; Madec, F.; Lampreave, F.

    2005-01-01

    In this work, apolipoprotein A-I (ApoA-I) was purified from pig sera. The responses of this protein after sterile inflammation and in animals infected with Actinobacillus pleuropneumoniae or Streptococcus suis were investigated. Decreases in the concentrations of ApoA-I, two to five times lower than the initial values, were observed at 2 to 4 days. It is concluded that ApoA-I is a negative acute-phase protein in pigs. PMID:15845530

  16. Macrophage metalloproteinases degrade high-density-lipoprotein-associated apolipoprotein A-I at both the N- and C-termini.

    PubMed Central

    Eberini, Ivano; Calabresi, Laura; Wait, Robin; Tedeschi, Gabriella; Pirillo, Angela; Puglisi, Lina; Sirtori, Cesare R; Gianazza, Elisabetta

    2002-01-01

    Atheromatous plaques contain various cell types, including macrophages, endothelial cells and smooth-muscle cells. To investigate the possible interactions between secreted matrix metalloproteinases and high-density lipoprotein (HDL) components, we tested the above cell types by culturing them for 24 h. HDL(3) (HDL subfractions with average sizes of between 8.44 nm for HDL(3A) and 7.62 nm for HDL(3C)) were then incubated in their cell-free conditioned media. Proteolytic degradation of apolipoprotein A-I was observed with macrophages, but not with endothelial-cell- or muscle-cell-conditioned supernatant. Absence of calcium or addition of EDTA to incubation media prevented all proteolytic processes. The identified apolipoprotein A-I fragments had sizes of 26, 22, 14 and 9 kDa. Two-dimensional electrophoresis and MS resolved the 26 and the 22 kDa components and identified peptides resulting from both N- and C-terminal cleavage of apolipoprotein A-I. The higher abundance of C- than N-terminally cleaved peptides agrees with data in the literature for a fully structured alpha-helix around Tyr(18) compared with an unstructured region around Gly(185) and Gly(186). The flexibility in the latter region of apolipoprotein A-I may explain its susceptibility to proteolysis. In our experimental set-up, HDL(3C) was more extensively degraded than the other HDL(3) subclasses (HDL(3A) and HDL(3B)). Proteolytic fragments produced by metalloproteinase action were shown by gel filtration and electrophoresis to be neither associated with lipids nor self-associated. PMID:11879189

  17. Serum apolipoprotein A-I is a novel prognostic indicator for non-metastatic nasopharyngeal carcinoma.

    PubMed

    Luo, Xiao-Lin; Zhong, Guang-Zheng; Hu, Li-Yang; Chen, Jie; Liang, Ying; Chen, Qiu-Yan; Liu, Qing; Rao, Hui-Lan; Chen, Kai-Lin; Cai, Qing-Qing

    2015-12-22

    We investigated the value of pretreatment serum apolipoprotein A-I (ApoA-I) in complementing TNM staging in the prognosis of non-metastatic nasopharyngeal carcinoma (NPC). We retrospectively reviewed 1196 newly diagnosed patients with non-metastatic NPC. Disease-specific survival (DSS), distant metastasis-free survival (DMFS), and locoregional recurrence-free survival (LRFS) rates were compared according to serum ApoA-I level. Multivariate analysis was performed to assess the prognostic value of serum ApoA-I. The 5-year DSS, DMFS, and LRFS rates for patients with elevated or decreased serum ApoA-I were 81.3% versus 69.3% (P < 0.001), 83.4% versus 67.4% (P < 0.001), and 80.9% versus 67.3% (P < 0.001), respectively. ApoA-I ≥ 1.025 g/L was an independent prognostic factor for superior DSS, DMFS, and LRFS in multivariate analysis. After stratification by clinical stage, serum ApoA-I remained a clinically and statistically significant predictor of prognosis. Our data suggest that the level of ApoA-I at diagnosis is a novel independent prognostic marker that could complement clinical staging for risk definition in non-metastatic NPC.

  18. Newly developed apolipoprotein A-I mimetic peptide promotes macrophage reverse cholesterol transport in vivo.

    PubMed

    Shimizu, Tomohiko; Tanigawa, Hiroyuki; Miura, Shin-ichiro; Kuwano, Takashi; Takata, Kohei; Suematsu, Yasunori; Imaizumi, Satoshi; Yahiro, Eiji; Zhang, Bo; Uehara, Yoshinari; Saku, Keijiro

    2015-08-01

    We elucidated the effect of newly developed Fukuoka Apolipoprotein A-I Mimetic Peptide (FAMP) on in vivo macrophage reverse cholesterol transport (RCT) and the underlying mechanisms. Cholesteryl ester transfer protein transgenic mice were divided into FAMP, and placebo control groups, and injected with FAMP or phosphate buffer saline intraperitoneally for 5 days. The FAMP group showed a significant decrease in plasma high-density lipoprotein cholesterol (HDL-C), and plasma from the FAMP group had an increased ability to promote ATP-binding cassette transporter A1 (ABCA1)-mediated cholesterol efflux from bone marrow macrophages ex vivo. Furthermore, mice were injected intraperitoneally with (3)H-cholesterol-labeled and cholesterol-loaded macrophages and monitored for the appearance of (3)H-tracer. The amount of (3)H-tracer excreted into feces over 48h in the FAMP group was significantly higher than that in the control group. (3)H-cholesterol ester (CE)-HDL was injected intravenously and (3)H-cholesterol in blood was counted. In the FAMP group, plasma (3)H-CE-HDL decreased rapidly, and treatment with FAMP markedly increased the fractional catabolic rate. The administration of FAMP promoted ABCA1-dependent efflux ex vivo, HDL turnover in vivo, and macrophage RCT in vivo despite reduced plasma HDL-C levels. FAMP might have atheroprotective potential. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  19. Matrix metalloproteinase 8 degrades apolipoprotein A-I and reduces its cholesterol efflux capacity.

    PubMed

    Salminen, Aino; Åström, Pirjo; Metso, Jari; Soliymani, Rabah; Salo, Tuula; Jauhiainen, Matti; Pussinen, Pirkko J; Sorsa, Timo

    2015-04-01

    Various cell types in atherosclerotic lesions express matrix metalloproteinase (MMP)-8. We investigated whether MMP-8 affects the structure and antiatherogenic function of apolipoprotein (apo) A-I, the main protein component of HDL particles. Furthermore, we studied serum lipid profiles and cholesterol efflux capacity in MMP-8-deficient mouse model. Incubation of apoA-I (28 kDa) with activated MMP-8 yielded 22 kDa and 25 kDa apoA-I fragments. Mass spectrometric analyses revealed that apoA-I was cleaved at its carboxyl-terminal part. Treatment of apoA-I and HDL with MMP-8 resulted in significant reduction (up to 84%, P < 0.001) in their ability to facilitate cholesterol efflux from cholesterol-loaded THP-1 macrophages. The cleavage of apoA-I by MMP-8 and the reduction in its cholesterol efflux capacity was inhibited by doxycycline. MMP-8-deficient mice had significantly lower serum triglyceride (TG) levels (P = 0.003) and larger HDL particles compared with wild-type (WT) mice. However, no differences were observed in the apoA-I levels or serum cholesterol efflux capacities between the mouse groups. Proteolytic modification of apoA-I by MMP-8 may impair the first steps of reverse cholesterol transport, leading to increased accumulation of cholesterol in the vessel walls. Eventually, inhibition of MMPs by doxycycline may reduce the risk for atherosclerotic vascular diseases.

  20. Thermal Stability of Apolipoprotein A-I in High-Density Lipoproteins by Molecular Dynamics

    PubMed Central

    Jones, Martin K.; Catte, Andrea; Patterson, James C.; Gu, Feifei; Chen, Jianguo; Li, Ling; Segrest, Jere P.

    2009-01-01

    Abstract Apolipoprotein (apo) A-I is an unusually flexible protein whose lipid-associated structure is poorly understood. Thermal denaturation, which is used to measure the global helix stability of high-density lipoprotein (HDL)-associated apoA-I, provides no information about local helix stability. Here we report the use of temperature jump molecular dynamics (MD) simulations to scan the per-residue helix stability of apoA-I in phospholipid-rich HDL. When three 20 ns MD simulations were performed at 500 K on each of two particles created by MD simulations at 310 K, bilayers remained intact but expanded by 40%, and total apoA-I helicity decreased from 95% to 72%. Of significance, the conformations of the overlapping N- and C-terminal domains of apoA-I in the particles were unusually mobile, exposing hydrocarbon regions of the phospholipid to solvent; a lack of buried interhelical salt bridges in the terminal domains correlated with increased mobility. Nondenaturing gradient gels show that 40% expansion of the phospholipid surface of 100:2 particles by addition of palmitoyloleoylphosphatidylcholine exceeds the threshold of particle stability. As a unifying hypothesis, we propose that the terminal domains of apoA-I are phospholipid concentration-sensitive molecular triggers for fusion/remodeling of HDL particles. Since HDL remodeling is necessary for cholesterol transport, our model for remodeling has substantial biomedical implications. PMID:19167289

  1. Tubulointerstitial nephritis is a dominant feature of hereditary apolipoprotein A-I amyloidosis.

    PubMed

    Gregorini, Gina; Izzi, Claudia; Ravani, Pietro; Obici, Laura; Dallera, Nadia; Del Barba, Andrea; Negrinelli, Alessandro; Tardanico, Regina; Nardi, Matilde; Biasi, Luciano; Scalvini, Tiziano; Merlini, Giampaolo; Scolari, Francesco

    2015-06-01

    Apolipoprotein A-I is the main protein of high-density lipoprotein particles, and is encoded by the APOA1 gene. Several APOA1 mutations have been found, either affecting the lecithin:cholesterol acyltransferase activity, determining familial HDL deficiency, or resulting in amyloid formation with prevalent deposits in the kidney and liver. Evaluation of familial tubulointerstitial nephritis in patients with the Leu75Pro APOA-I amyloidosis mutation resulted in the identification of 253 carriers belonging to 50 families from Brescia, Italy. A total of 219 mutation carriers underwent clinical, laboratory, and instrumental tests. Of these, 62% had renal, hepatic, and testicular disease; 38% were asymptomatic. The disease showed an age-dependent penetrance. Tubulointerstitial nephritis was diagnosed in 49% of the carriers, 13% of whom progressed to kidney failure requiring dialysis. Hepatic involvement with elevation of cholestasis indices was diagnosed in 30% of the carriers, 38% of whom developed portal hypertension. Impaired spermatogenesis and hypogonadism was found in 68% of male carriers. The cholesterol levels were lower than normal in 80% of the mutation carriers. Thus, tubulointerstitial nephritis was highly prevalent in this large series of patients with Leu75Pro apoA-I amyloidosis. Persistent elevation of alkaline phosphatase, reduced HDL cholesterol plasma levels, and hypogonadism in men are key diagnostic features of this form of amyloidosis.

  2. Apolipoprotein A-I and B and Subjective Global Assessment relationship can reflect lipid defects in diabetic retinopathy.

    PubMed

    Sharma, Yashodhara; Saxena, Sandeep; Mishra, Arvind; Saxena, Anita; Natu, Shankar Madhav

    2017-01-01

    Elevated lipid levels increase complications of diabetic retinopathy (DR). Uncontrolled diabetes increases these complications and causes unintentional weight loss, indicating an apparently normal body mass index (BMI). Thus, it is easy to assume that patients with DR and a normal BMI have optimal lipid status. Apolipoprotein (Apo) A-I and Apo B levels differentially indicate serum lipid status in DR. Subjective Global Assessment (SGA) scores are associated with DR status. If SGA scores and serum Apo A-I and B levels are found to be interrelated, their relationship can reflect lipid defects in patients with DR despite apparently normal BMI. The aim of the present study was to investigate the possible relationship between serum Apo A-I and B levels and SGA scores of patients with DR. This was a case-control study conducted from November 2011 to April 2014. Serum Apo A-I and B levels and SGA scores were calculated for 40 healthy controls, 48 individuals without DR, 49 nonproliferative DR cases, and 48 proliferative DR cases. Pearson's correlation analysis was applied between Apo A-I, Apo B, Apo B/Apo A-I ratio, and SGA scores. Negative correlation was observed between serum Apo A-I level (r = -0.567, P < 0.001) and positive correlation between serum Apo B level (r = 0.451, P < 0.001) and Apo B/Apo A-I ratio (r = 0.597, P < 0.001) with escalating SGA scores. To our knowledge, this is the first study to report a novel correlation between serum Apo A-I, Apo B and Apo B/Apo A-I ratio and SGA scores. SGA scores can help predict lipid abnormalities in patients with DR even when they have an apparently normal BMI. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Lipid-free Apolipoprotein A-I Structure: Insights into HDL Formation and Atherosclerosis Development

    PubMed Central

    Mei, Xiaohu; Atkinson, David

    2015-01-01

    Apolipoprotein A-I is the major protein in high-density lipoprotein (HDL) and plays an important role during the process of reverse cholesterol transport (RCT). Knowledge of the high-resolution structure of full-length apoA-I is vital for a molecular understanding of the function of HDL at the various steps of the RCT pathway. Due to the flexible nature of apoA-I and aggregation properties, the structure of full-length lipid-free apoA-I has evaded description for over three decades. Sequence analysis of apoA-I suggested that the amphipathic α-helix is the structural motif of exchangeable apolipoprotein, and NMR, X-ray and MD simulation studies have confirmed this. Different laboratories have used different methods to probe the secondary structure distribution and organization of both the lipid-free and lipid-bound apoA-I structure. Mutation analysis, synthetic peptide models, surface chemistry and crystal structures have converged on the lipid-free apoA-I domain structure and function: the N-terminal domain [1–184] forms a helix bundle while the C-terminal domain [185–243] mostly lacks defined structure and is responsible for initiating lipid-binding, aggregation and is also involved in cholesterol efflux. The first 43 residues of apoA-I are essential to stabilize the lipid-free structure. In addition, the crystal structure of C-terminally truncated apoA-I suggests a monomer-dimer conversation mechanism mediated through helix 5 reorganization and dimerization during the formation of HDL. Based on previous research, we have proposed a structural model for full-length monomeric apoA-I in solution and updated the HDL formation mechanism through three intermediate states. Mapping the known natural mutations on the full-length monomeric apoA-I model provides insight into atherosclerosis development through disruption of the N-terminal helix bundle or deletion of the C-terminal lipid-binding domain. PMID:26048453

  4. Increased Binding of Apolipoproteins A-I and E4 to Triglyceride-Rich Lipoproteins is linked to Induction of Hypertriglyceridemia.

    PubMed

    Gorshkova, Irina N; Atkinson, David

    2017-01-01

    Hypertriglyceridemia (HTG) is an independent factor of atherosclerotic cardiovascular disease and a hallmark of many metabolic disorders. However, the molecular etiology of HTG is still largely unknown. In mice, severe HTG may be induced by expression of specific mutants of apolipoprotein (apo) A-I or wild type (WT) apoE4. Expression of a certain apoE4 mutant results in mild HTG, while expression of another apoE4 mutant or WT apoA-I results in normal plasma triglyceride (TG) levels. Biophysical studies of the apoA-I and apoE4 forms associated with HTG help better understand the molecular mechanisms of induction of HTG by these proteins. The studies show that the apoA-I and apoE4 forms that induce HTG have a destabilized and more loosely folded conformation in solution than their counterparts not associated with HTG. Disruption of the protein salt bridge networks by the mutations is likely responsible for the observed structural changes. Each apoA-I and apoE4 form that induced HTG show enhanced binding to model TG-rich particles. HTG appeared to positively correlate with the apolipoprotein ability to bind to TG-rich particles. This implies that in vivo, the conformational changes in the apolipoproteins that induce HTG facilitate their binding to plasma TG-rich lipoproteins. We discuss metabolic pathways leading to the development of HTG that may result from enhanced binding of the apolipoproteins to TG-rich lipoproteins in circulation. While various factors may be involved in the development of HTG in humans, it is possible that structural alterations that increase affinity of apolipoproteins to TG-rich lipoproteins may contribute to some cases of this disorder.

  5. Surface-induced assembly of apolipoprotein A-I: Implications for symmetry-driven non-cooperative clustering

    NASA Astrophysics Data System (ADS)

    Winford, Sidney; Tobin, Moriah; Gross, Eitan

    2012-03-01

    In condensed matter physics the geometry of a crystal is determined by the mechanism of condensation. In biology, the link between clustering mechanisms and the shape of a protein crystal is not well defined. To gain more insight into the problem, we studied clustering of apolipoprotein A-I (apo A-I) on a solid surface using AFM. The amyloidogenic protein apo A-I is the main protein component of high density lipoprotein and thus reduces the risk of atherosclerosis. We found that apo A-I clustered to form nano-scale, symmetrical clusters on mica. Statistical analysis of size distribution for several thousand clusters suggested that the clustering reaction followed a non-cooperative kinetic scheme characterized by a single equilibrium constant of 0.92·106 M-1 and a change in free energy (ΔG) of -0.03 kJ mole-1/residue. This is close to ΔG of-0.04 kJ mole-1/residue for apo A-I binding to phospholipid membrane; and 30-fold smaller than ΔG for β-amyloid formation by apo A-I. The high symmetry of the clusters is consistent with an isotropic diffusion coefficient of protein monomers on the surface of the substrate. This previously unrecognized link between protein clustering mechanism and the symmetry of the growth pattern may have important implications in medicine, pharmaceutics and polymer science.

  6. Formation of stable nanodiscs by bihelical apolipoprotein A-I mimetic peptide.

    PubMed

    Kariyazono, Hirokazu; Nadai, Ryo; Miyajima, Rin; Takechi-Haraya, Yuki; Baba, Teruhiko; Shigenaga, Akira; Okuhira, Keiichiro; Otaka, Akira; Saito, Hiroyuki

    2016-02-01

    Nanodiscs are composed of scaffold protein or peptide such as apolipoprotein A-I (apoA-I) and phospholipids. Although peptide-based nanodiscs have an advantage to modulate the size of nanodiscs by changing phospholipid/peptide ratios, they are usually less stable than apoA-I-based nanodiscs. In this study, we designed a novel nanodisc scaffold peptide (NSP) that has proline-punctuated bihelical amphipathic structure based on apoA-I mimetic peptides. NSP formed α-helical structure on 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) nanodiscs prepared by cholate dialysis method. Dynamic light scattering measurements demonstrated that diameters of NSP nanodiscs vary depending upon POPC/NSP ratios. Comparison of thermal unfolding of nanodiscs monitored by circular dichroism measurements demonstrated that NSP forms much more stable nanodiscs with POPC than monohelical peptide, 4F, exhibiting comparable stability to apoA-I-POPC nanodiscs. Intrinsic Trp fluorescence measurements showed that Trp residues of NSP exhibit more hydrophobic environment than that of 4 F on nanodiscs, suggesting the stronger interaction of NSP with phospholipids. Thus, the bihelical structure of NSP appears to increase the stability of nanodiscs because of the enhanced interaction of peptides with phospholipids. In addition, NSP as well as 4F spontaneously solubilized POPC vesicles into nanodiscs without using detergent. These results indicate that bihelical NSP forms nanodiscs with comparable stability to apoA-I and has an ability to control the size of nanodiscs simply by changing phospholipid/peptide ratios. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  7. Apolipoprotein A-I mimetic peptide reverses impaired arterial healing after injury by reducing oxidative stress.

    PubMed

    Rosenbaum, Michael A; Chaudhuri, Pinaki; Abelson, Benjamin; Cross, Brandy N; Graham, Linda M

    2015-08-01

    Endothelial cell (EC) migration is essential for healing of arterial injuries caused by angioplasty, but a high cholesterol diet inhibits endothelial repair. In vivo studies suggest that apolipoprotein A-I (apoA-I), the major protein constituent of HDL, is essential for normal healing of arterial injuries. ApoA-I mimetics, including 4F, have been designed to mimic the amphipathic portion of the apoA-I molecule. This study was undertaken to determine if 4F improves endothelial migration and healing. A razor scrape assay was used to analyze the effect of 4F on EC migration in vitro. Endothelial healing in vivo was assessed following electrical injury of carotid arteries in mice. Markers of oxidative stress were also examined. Lipid oxidation products inhibited EC migration in vitro, but preincubation with L-4F preserved EC migration. Endothelial healing of carotid arterial injuries in mice on a high cholesterol diet was delayed compared with mice on a chow diet with 27.8% vs. 48.2% healing, respectively, at 5 days. Administration of D-4F improved endothelial healing in mice on a high cholesterol diet to 43.4%. D-4F administration had no effect on lipid levels but decreased markers of oxidation. In vivo, there was a significant inverse correlation between endothelial healing and plasma markers of oxidative stress. These studies suggested that an apoA-I mimetic can improve endothelial healing of arterial injuries by decreasing oxidative stress. Published by Elsevier Ireland Ltd.

  8. Abdominal obesity with hypertriglyceridaemia, lipoprotein(a) and apolipoprotein A-I determine marked cardiometabolic risk.

    PubMed

    Onat, Altan; Can, Günay; Örnek, Ender; Sansoy, Vedat; Aydın, Mesut; Yüksel, Hüsniye

    2013-11-01

    Risks for coronary heart disease (CHD) and diabetes (T2DM) of the 'hypertriglyceridemic waist' phenotype (HtgW) warrant further investigation. We studied this issue and whether partial proinflammatory conversion of apolipoprotein (apo) A-I by lipoprotein(a) [Lp(a)] is a codeterminant. In a population-based prospective study, 1328 Turkish adults were analysed in four groups by the presence of abdominal obesity and elevated triglycerides (Htg). LDL-cholesterol levels, significantly elevated in isolated Htg, were lower in HtgW, yet significantly higher apoB and complement C3 values existed in women with HtgW in whom also the lowest Lp(a) values prevailed. Lp(a) was linearly associated, more strongly in HtgW than in the remaining groups, with apoB and, in women inversely, with gamma-glutamyltransferase. Incident HtgW was predicted, not in men, but in women inversely by Lp(a) (OR 0.80 [95%CI 0.65; 0.97]), regardless of adjustment for relevant confounders. After adjustment for conventional risk factors, HtgW (OR 2.84) and high apoA-I/HDL-C ratio (OR 1.50) were significantly and additively associated with combined prevalent and incident CHD risk. High apoA-I and low HDL-cholesterol levels interacted therein in women. Type-2 diabetes was strongly predicted by HtgW, mediated in men by high apoA-I/HDL-C ratio. HtgW is associated with excess inflammatory markers, is predicted in women paradoxically by lower circulating Lp(a) and is associated in both sexes with marked excess cardiometabolic risk to which high apoA-I/HDL-C ratio contributes additively. These findings are consistent in women with apoA-I being oxidized via aggregation to Lp(a). © 2013 Stichting European Society for Clinical Investigation Journal Foundation. Published by John Wiley & Sons Ltd.

  9. Surface behavior of apolipoprotein A-I and its deletion mutants at model lipoprotein interfaces

    PubMed Central

    Wang, Libo; Mei, Xiaohu; Atkinson, David; Small, Donald M.

    2014-01-01

    Apolipoprotein A-I (apoA-I) has a great conformational flexibility to exist in lipid-free, lipid-poor, and lipid-bound states during lipid metabolism. To address the lipid binding and the dynamic desorption behavior of apoA-I at lipoprotein surfaces, apoA-I, Δ(185-243)apoA-I, and Δ(1-59)(185-243)apoA-I were studied at triolein/water and phosphatidylcholine/triolein/water interfaces with special attention to surface pressure. All three proteins are surface active to both interfaces lowering the interfacial tension and thus increasing the surface pressure to modify the interfaces. Δ(185-243)apoA-I adsorbs much more slowly and lowers the interfacial tension less than full-length apoA-I, confirming that the C-terminal domain (residues 185-243) initiates the lipid binding. Δ(1-59)(185-243)apoA-I binds more rapidly and lowers the interfacial tension more than Δ(185-243)apoA-I, suggesting that destabilizing the N-terminal α-helical bundle (residues 1-185) restores lipid binding. The three proteins desorb from both interfaces at different surface pressures revealing that different domains of apoA-I possess different lipid affinity. Δ(1-59)(185-243)apoA-I desorbs at lower pressures compared with apoA-I and Δ(185-243)apoA-I indicating that it is missing a strong lipid association motif. We propose that during lipoprotein remodeling, surface pressure mediates the adsorption and partial or full desorption of apoA-I allowing it to exchange among different lipoproteins and adopt various conformations to facilitate its multiple functions. PMID:24308948

  10. Apolipoprotein A-I mimetic peptide treatment inhibits inflammatory responses and improves survival in septic rats

    PubMed Central

    Zhang, Zhenghao; Datta, Geeta; Zhang, Yun; Miller, Andrew P.; Mochon, Paulina; Chen, Yiu-Fai; Chatham, John; Anantharamaiah, G. M.; White, C. Roger

    2009-01-01

    Systemic inflammation induces a multiple organ dysfunction syndrome that contributes to morbidity and mortality in septic patients. Since increasing plasma apolipoprotein A-I (apoA-I) and HDL may reduce the complications of sepsis, we tested the hypothesis that the apoA-I mimetic peptide 4F confers similar protective effects in rats undergoing cecal ligation and puncture (CLP) injury. Male Sprague-Dawley rats were randomized to undergo CLP or sham surgery. IL-6 levels were significantly elevated in plasma by 6 h after CLP surgery compared with shams. In subsequent studies, CLP rats were further subdivided to receive vehicle or 4F (10 mg/kg) by intraperitoneal injection, 6 h after sepsis induction. Sham-operated rats received saline. Echocardiographic studies showed a reduction in left ventricular end-diastolic volume, stroke volume, and cardiac output (CO) 24 h after CLP surgery. These changes were associated with reduced blood volume and left ventricular filling pressure. 4F treatment improved blood volume status, increased CO, and reduced plasma IL-6 in CLP rats. Total cholesterol (TC) and HDL were 79 ± 5 and 61 ± 4 mg/dl, respectively, in sham rats. TC was significantly reduced in CLP rats (54 ± 3 mg/dl) due to a reduction in HDL (26 ± 3 mg/dl). 4F administration to CLP rats attenuated the reduction in TC (69 ± 4 mg/dl) and HDL (41 ± 3 mg/dl) and prevented sepsis-induced changes in HDL protein composition. Increased plasma HDL in 4F-treated CLP rats was associated with an improvement in CO and reduced mortality. It is proposed that protective effects of 4F are related to its ability to prevent the sepsis-induced reduction in plasma HDL. PMID:19561306

  11. Apolipoprotein A-I Helsinki promotes intracellular acyl-CoA cholesterol acyltransferase (ACAT) protein accumulation.

    PubMed

    Toledo, Juan D; Garda, Horacio A; Cabaleiro, Laura V; Cuellar, Angela; Pellon-Maison, Magali; Gonzalez-Baro, Maria R; Gonzalez, Marina C

    2013-05-01

    Reverse cholesterol transport is a process of high antiatherogenic relevance in which apolipoprotein AI (apoA-I) plays an important role. The interaction of apoA-I with peripheral cells produces through mechanisms that are still poorly understood the mobilization of intracellular cholesterol depots toward plasma membrane. In macrophages, these mechanisms seem to be related to the modulation of the activity of acyl-CoA cholesterol acyltransferase (ACAT), the enzyme responsible for the intracellular cholesterol ester biosynthesis that is stored in lipid droplets. The activation of ACAT and the accumulation of lipid droplets play a key role in the transformation of macrophages into foam cells, leading to the formation of atheroma or atherosclerotic plaque. ApoA-I Helsinki (or ∆K107) is a natural apoA-I variant with a lysine deletion in the central protein region, carriers of which have increased atherosclerosis risk. We herein show that treatment of cultured RAW macrophages or CHOK1 cells with ∆K107, but not with wild-type apoA-I or a variant containing a similar deletion at the C-terminal region (∆K226), lead to a marked increase (more than 10 times) in the intracellular ACAT1 protein level as detected by western blot analysis. However, we could only detect a slight increase in cholesteryl ester produced by ∆K107 mainly when Chol loading was supplied by low-density lipoprotein (LDL). Although a similar choline-phospholipid efflux is evoked by these apoA-I variants, the change in phosphatidylcholine/sphyngomyelin distribution produced by wild-type apoA-I is not observed with either ∆K107 or ∆K226.

  12. Myeloperoxidase-mediated Methionine Oxidation Promotes an Amyloidogenic Outcome for Apolipoprotein A-I*

    PubMed Central

    Chan, Gary K. L.; Witkowski, Andrzej; Gantz, Donald L.; Zhang, Tianqi O.; Zanni, Martin T.; Jayaraman, Shobini; Cavigiolio, Giorgio

    2015-01-01

    High plasma levels of apolipoprotein A-I (apoA-I) correlate with cardiovascular health, whereas dysfunctional apoA-I is a cause of atherosclerosis. In the atherosclerotic plaques, amyloid deposition increases with aging. Notably, apoA-I is the main component of these amyloids. Recent studies identified high levels of oxidized lipid-free apoA-I in atherosclerotic plaques. Likely, myeloperoxidase (MPO) secreted by activated macrophages in atherosclerotic lesions is the promoter of such apoA-I oxidation. We hypothesized that apoA-I oxidation by MPO levels similar to those present in the artery walls in atherosclerosis can promote apoA-I structural changes and amyloid fibril formation. ApoA-I was exposed to exhaustive chemical (H2O2) oxidation or physiological levels of enzymatic (MPO) oxidation and incubated at 37 °C and pH 6.0 to induce fibril formation. Both chemically and enzymatically oxidized apoA-I produced fibrillar amyloids after a few hours of incubation. The amyloid fibrils were composed of full-length apoA-I with differential oxidation of the three methionines. Met to Leu apoA-I variants were used to establish the predominant role of oxidation of Met-86 and Met-148 in the fibril formation process. Importantly, a small amount of preformed apoA-I fibrils was able to seed amyloid formation in oxidized apoA-I at pH 7.0. In contrast to hereditary amyloidosis, wherein specific mutations of apoA-I cause protein destabilization and amyloid deposition, oxidative conditions similar to those promoted by local inflammation in atherosclerosis are sufficient to transform full-length wild-type apoA-I into an amyloidogenic protein. Thus, MPO-mediated oxidation may be implicated in the mechanism that leads to amyloid deposition in the atherosclerotic plaques in vivo. PMID:25759391

  13. High-density lipoprotein apolipoprotein A-I kinetics in obesity.

    PubMed

    Ooi, Esther M M; Watts, Gerald F; Farvid, Maryam S; Chan, Dick C; Allen, Michael C; Zilko, Simon R; Barrett, P Hugh R

    2005-06-01

    Low plasma concentrations of high-density lipoprotein (HDL)-cholesterol and apolipoprotein A-I (apoA-I) are independent predictors of coronary artery disease and are often associated with obesity and the metabolic syndrome. However, the underlying kinetic determinants of HDL metabolism are not well understood. We pooled data from 13 stable isotope studies to investigate the kinetic determinants of apoA-I concentrations in lean and overweight-obese individuals. We also examined the associations of HDL kinetics with age, sex, BMI, fasting plasma glucose, fasting insulin, Homeostasis Model Assessment score, and concentrations of apoA-I, triglycerides, HDL-cholesterol and low-density lipoprotein-cholesterol. Compared with lean individuals, overweight-obese individuals had significantly higher HDL apoA-I fractional catabolic rate (0.21+/-0.01 vs. 0.33+/-0.01 pools/d; p<0.001) and production rate (PR; 11.3+/-4.4 vs. 15.8+/-2.77 mg/kg per day; p=0.001). In the lean group, HDL apoA-I PR was significantly associated with apoA-I concentration (r=0.455, p=0.004), whereas in the overweight-obese group, both HDL apoA-I fractional catabolic rate (r=-0.396, p=0.050) and HDL apoA-I PR (r=0.399, p=0.048) were significantly associated with apoA-I concentration. After adjustment for fasting insulin or Homeostasis Model Assessment score, HDL apoA-I PR was an independent predictor of apoA-I concentration. In overweight-obese subjects, hypercatabolism of apoA-I is paralleled by an increased production of apoA-I, with HDL apoA-I PR being the stronger determinant of apoA-I concentration. This could have therapeutic implications for the management of dyslipidemia in individuals with low plasma HDL-cholesterol.

  14. Apolipoprotein A-I Is a Potential Mediator of Remote Ischemic Preconditioning

    PubMed Central

    Hibert, Pierre; Prunier-Mirebeau, Delphine; Beseme, Olivia; Chwastyniak, Maggy; Tamareille, Sophie; Lamon, Delphine; Furber, Alain; Pinet, Florence; Prunier, Fabrice

    2013-01-01

    Background Remote ischemic preconditioning (RIPC) has emerged as an attractive strategy in clinical settings. Despite convincing evidence of the critical role played by circulating humoral mediators, their actual identities remain unknown. In this study, we aimed to identify RIPC-induced humoral mediators using a proteomic approach. Methods and Results Rats were exposed to 10-min limb ischemia followed by 5- (RIPC 5′) or 10-min (RIPC 10′) reperfusion prior to blood sampling. The control group only underwent blood sampling. Plasma samples were analyzed using surface-enhanced laser desorption and ionization - time of flight - mass spectrometry (SELDI-TOF-MS). Three protein peaks were selected for their significant increase in RIPC 10′. They were identified and confirmed as apolipoprotein A-I (ApoA-I). Additional rats were exposed to myocardial ischemia-reperfusion (I/R) and assigned to one of the following groups RIPC+myocardial infarction (MI) (10-min limb ischemia followed by 10-min reperfusion initiated 20 minutes prior to myocardial I/R), ApoA-I+MI (10 mg/kg ApoA-I injection 10 minutes before myocardial I/R), and MI (no further intervention). In comparison with untreated MI rats, RIPC reduced infarct size (52.2±3.7% in RIPC+MI vs. 64.9±2.6% in MI; p<0.05). Similarly, ApoA-I injection decreased infarct size (50.9±3.8%; p<0.05 vs. MI). Conclusions RIPC was associated with a plasmatic increase in ApoA-I. Furthermore, ApoA-I injection before myocardial I/R recapitulated the cardioprotection offered by RIPC in rats. This data suggests that ApoA-I may be a protective blood-borne factor involved in the RIPC mechanism. PMID:24155931

  15. Isolation of human apolipoprotein E by chromatofocusing.

    PubMed

    Weisweiler, P; Schwandt, P

    1982-09-01

    Human prolipoprotein E is implicated in the transport of serum cholesterol and the binding of lipoproteins to cell receptors. Further investigations on this apolipoprotein would be facilitated by improved purification methods. We prepared human apo E by the combination of high performance gel filtration and chromatofocusing from serum very low density lipoproteins. Chromatofocusing was performed with a pH gradient from 7 to 4. Apo E contained all isoforms, but was homogeneous in SDS-polyacrylamide gel electrophoresis and in double immunodiffusion against a monospecific antiserum. The reported purification method allows a rapid and simple preparation of large amounts of apo E.

  16. Apolipoprotein A-I mimetic peptide 4F rescues pulmonary hypertension by inducing microRNA-193-3p.

    PubMed

    Sharma, Salil; Umar, Soban; Potus, Francois; Iorga, Andrea; Wong, Gabriel; Meriwether, David; Breuils-Bonnet, Sandra; Mai, Denise; Navab, Kaveh; Ross, David; Navab, Mohamad; Provencher, Steeve; Fogelman, Alan M; Bonnet, Sébastien; Reddy, Srinivasa T; Eghbali, Mansoureh

    2014-08-26

    Pulmonary arterial hypertension is a chronic lung disease associated with severe pulmonary vascular changes. A pathogenic role of oxidized lipids such as hydroxyeicosatetraenoic and hydroxyoctadecadienoic acids is well established in vascular disease. Apolipoprotein A-I mimetic peptides, including 4F, have been reported to reduce levels of these oxidized lipids and improve vascular disease. However, the role of oxidized lipids in the progression of pulmonary arterial hypertension and the therapeutic action of 4F in pulmonary arterial hypertension are not well established. We studied 2 different rodent models of pulmonary hypertension (PH): a monocrotaline rat model and a hypoxia mouse model. Plasma levels of hydroxyeicosatetraenoic and hydroxyoctadecadienoic acids were significantly elevated in PH. 4F treatment reduced these levels and rescued preexisting PH in both models. MicroRNA analysis revealed that microRNA-193-3p (miR193) was significantly downregulated in the lung tissue and serum from both patients with pulmonary arterial hypertension and rodents with PH. In vivo miR193 overexpression in the lungs rescued preexisting PH and resulted in downregulation of lipoxygenases and insulin-like growth factor-1 receptor. 4F restored PH-induced miR193 expression via transcription factor retinoid X receptor α. These studies establish the importance of microRNAs as downstream effectors of an apolipoprotein A-I mimetic peptide in the rescue of PH and suggest that treatment with apolipoprotein A-I mimetic peptides or miR193 may have therapeutic value. © 2014 American Heart Association, Inc.

  17. High 99mTc-DPD myocardial uptake in a patient with apolipoprotein AI-related amyloidotic cardiomyopathy.

    PubMed

    Quarta, Candida Cristina; Obici, Laura; Guidalotti, Pier Luigi; Pieroni, Maurizio; Longhi, Simone; Perlini, Stefano; Verga, Laura; Merlini, Giampaolo; Rapezzi, Claudio

    2013-03-01

    Amyloidotic cardiomyopathy is still a widely underdiagnosed condition that usually requires endomyocardial biopsy (EMB) for a definite diagnosis. 99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid (99mTc-DPD) has proven highly sensitive for detecting amyloidotic cardiomyopathy due to transthyretin-related amyloid deposition. Herein we report the first description of the (99mTc-DPD scintigraphy profile in a patient with suspected amyloidotic cardiomyopathy and a final EMB- and genetically-proven diagnosis of familial apolipoprotein AI amyloidosis due to Leu174Ser variant.

  18. Myeloperoxidase-mediated Methionine Oxidation Promotes an Amyloidogenic Outcome for Apolipoprotein A-I.

    PubMed

    Chan, Gary K L; Witkowski, Andrzej; Gantz, Donald L; Zhang, Tianqi O; Zanni, Martin T; Jayaraman, Shobini; Cavigiolio, Giorgio

    2015-04-24

    High plasma levels of apolipoprotein A-I (apoA-I) correlate with cardiovascular health, whereas dysfunctional apoA-I is a cause of atherosclerosis. In the atherosclerotic plaques, amyloid deposition increases with aging. Notably, apoA-I is the main component of these amyloids. Recent studies identified high levels of oxidized lipid-free apoA-I in atherosclerotic plaques. Likely, myeloperoxidase (MPO) secreted by activated macrophages in atherosclerotic lesions is the promoter of such apoA-I oxidation. We hypothesized that apoA-I oxidation by MPO levels similar to those present in the artery walls in atherosclerosis can promote apoA-I structural changes and amyloid fibril formation. ApoA-I was exposed to exhaustive chemical (H2O2) oxidation or physiological levels of enzymatic (MPO) oxidation and incubated at 37 °C and pH 6.0 to induce fibril formation. Both chemically and enzymatically oxidized apoA-I produced fibrillar amyloids after a few hours of incubation. The amyloid fibrils were composed of full-length apoA-I with differential oxidation of the three methionines. Met to Leu apoA-I variants were used to establish the predominant role of oxidation of Met-86 and Met-148 in the fibril formation process. Importantly, a small amount of preformed apoA-I fibrils was able to seed amyloid formation in oxidized apoA-I at pH 7.0. In contrast to hereditary amyloidosis, wherein specific mutations of apoA-I cause protein destabilization and amyloid deposition, oxidative conditions similar to those promoted by local inflammation in atherosclerosis are sufficient to transform full-length wild-type apoA-I into an amyloidogenic protein. Thus, MPO-mediated oxidation may be implicated in the mechanism that leads to amyloid deposition in the atherosclerotic plaques in vivo. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Blood serum apolipoproteins B and A-I in females suffering from rheumatic heart valve disease.

    PubMed

    Stakisaitis, Donatas; Maksvytis, Arūnas; Salcius, Kestutis; Benetis, Rimantas

    2004-01-01

    With the aim to check whether the atherogenic factors are involved in the mechanisms of valve fibrosis, we have studied the blood serum concentrations of apolipoproteins (apo) A-I and B concentration in patients suffering from rheumatic heart valve fibrosis. The quantities of apoA-I and apoB in the blood serum were tested by the ELISA method. Concentration of apoA-I and B in the blood serum was determined in rheumatic females with replacement of the damaged valves: after aortic valve operation (n=11; mean age 43.3+/-3.6 years) and after mitral valve operation (n=29; 41.3+/-4.1). The results obtained for rheumatic patients were compared with the data on age-matched healthy females (n=43; 39.5+/-5.2 years). Significantly lower apoA-I level in the blood serum of all patients suffering from rheumatic heart valve disease was determined as compared with controls: in the pooled group of patients (1.02+/-0.22 vs 1.23+/-0.23 g/l, P<0.001), in women after aortic valve replacement (0.98+/-0.21 vs. 1.23+/-0.23 g/l, P<0.005), and in women after mitral valve surgery (1.03+/-0.23 g/l vs 1.23+/-0.23 g/l, P<0.005). The apoB level in patients suffering from rheumatic heart valve disease did not differ from that of controls. The apoB/apoA-I ratio for patients with valve fibrotic damage was significantly higher as compared to controls in all groups (P<0.02). The increase of apoB/apoA-I ratio in patients with rheumatic valve fibrosis was caused by lower apoA-I levels in blood serum. The obtained results indicate that decreased apoA-I levels in blood serum can be indicative of valve fibrosis in rheumatic heart valve disease patients.

  20. Apolipoprotein A-I gene transfer exerts immunomodulatory effects and reduces vascular inflammation and fibrosis in ob/ob mice.

    PubMed

    Spillmann, Frank; De Geest, Bart; Muthuramu, Ilayaraja; Amin, Ruhul; Miteva, Kapka; Pieske, Burkert; Tschöpe, Carsten; Van Linthout, Sophie

    2016-01-01

    Obesity is associated with vascular inflammation, fibrosis and reduced high-density lipoproteins (HDL)-cholesterol. We aimed to investigate whether adenoviral gene transfer with human apolipoprotein (apo) A-I (Ad.A-I), the main apo of HDL, could exert immunomodulatory effects and counteract vascular inflammation and fibrosis in ob/ob mice. Ad.A-I transfer was performed in 8 weeks (w) old ob/ob mice, which were sacrificed 7 w later. The aorta was excised for mRNA analysis and the spleen for splenocyte isolation for subsequent flow cytometry and co-culture with murine fibroblasts. HDL was added to mononuclear cells (MNC) and fibroblasts to assess their impact on adhesion capacity and collagen deposition, respectively. Ad.A-I led to a 1.8-fold (p < 0.05) increase in HDL-cholesterol versus control ob/ob mice at the day of sacrifice, which was paralleled by a decrease in aortic TNF-α and VCAM-1 mRNA expression. Pre-culture of MNC with HDL decreased their adhesion to TNF-α-activated HAEC. Ad.A-I exerted immunomodulatory effects as evidenced by a downregulation of aortic NOD2 and NLRP3 mRNA expression and by a 12 %, 6.9 %, and 15 % decrease of the induced proliferation/activity of total splenic MNC, CD4+, and CD8+ cells in ob/ob Ad.A-I versus control ob/ob mice, respectively (p < 0.05). Ad.A-I further reduced aortic collagen I and III mRNA expression by 62 % and 66 %, respectively (p < 0.0005), and abrogated the potential of ob/ob splenocytes to induce the collagen content in murine fibroblasts upon co-culture. Finally, HDL decreased the TGF-ß1-induced collagen deposition of murine fibroblasts in vitro. Apo A-I transfer counteracts vascular inflammation and fibrosis in ob/ob mice.

  1. A case report of hereditary apolipoprotein A-I amyloidosis associated with a novel APOA1 mutation and variable phenotype.

    PubMed

    Tougaard, Birgitte G; Pedersen, Katja Venborg; Krag, Søren Rasmus; Gilbertson, Janet A; Rowczenio, Dorota; Gillmore, Julian D; Birn, Henrik

    2016-09-01

    Apolipoprotein A-I (apo A-I) amyloidosis is a non-AL, non-AA, and non-transthyretin type of amyloidosis associated with mutations in the APOA1 gene inherited in an autosomal dominant fashion. It is a form of systemic amyloidosis, but at presentation, can also mimic localized amyloidosis. The renal presentation generally involves interstitial and medullary deposition of apo A-I amyloid protein. We describe the identification of apo A-I amyloidosis by mass spectrometry in a 52-year old male, with no family history of amyloidosis, presenting with nephrotic syndrome and associated with heterozygosity for a novel APOA1 mutation (c.220 T > A) which encodes the known amyloidogenic Trp50Arg variant. Renal amyloid deposits in this case were confined to the glomeruli alone, and the patient developed progressive renal impairment. One year after diagnosis, the patient had a successful kidney transplant from an unrelated donor. Pathogenic mutations in the APOA1 gene are generally associated with symptoms of amyloidosis. In this family however, genotyping of family members identified several unaffected carriers suggesting a variable disease penetrance, which has not been described before in this form of amyloidosis and has implications when counselling those with APOA1 mutations. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  2. Apolipoprotein A-I configuration and cell cholesterol efflux activity of discoidal lipoproteins depend on the reconstitution process.

    PubMed

    Cuellar, Luz Ángela; Prieto, Eduardo Daniel; Cabaleiro, Laura Virginia; Garda, Horacio Alberto

    2014-01-01

    Discoidal high-density lipoproteins (D-HDL) are critical intermediates in reverse cholesterol transport. Most of the present knowledge of D-HDL is based on studies with reconstituted lipoprotein complexes of apolipoprotein A-I (apoA-I) obtained by cholate dialysis (CD). D-HDL can also be generated by the direct microsolubilization (DM) of phospholipid vesicles at the gel/fluid phase transition temperature, a process mechanistically similar to the "in vivo" apoAI lipidation via ABCA1. We compared the apoA-I configuration in D-HDL reconstituted with dimyristoylphosphatidylcholine by both procedures using fluorescence resonance energy transfer measurements with apoA-I tryptophan mutants and fluorescently labeled cysteine mutants. Results indicate that apoA-I configuration in D-HDL depends on the reconstitution process and are consistent with a "double belt" molecular arrangement with different helix registry. As reported by others, a configuration with juxtaposition of helices 5 of each apoAI monomer (5/5 registry) predominates in D-HDL obtained by CD. However, a configuration with helix 5 of one monomer juxtaposed with helix 2 of the other (5/2 registry) would predominate in D-HDL generated by DM. Moreover, we also show that the kinetics of cholesterol efflux from macrophage cultures depends on the reconstitution process, suggesting that apoAI configuration is important for this HDL function. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Monoclonal antibodies to human apolipoproteins: application to the study of high density lipoprotein subpopulations.

    PubMed

    Bustos, P; Ulloa, N; Calvo, C; Muller, D; Durán, D; Martínez, J; Salazar, L; Quiroga, A

    2000-09-01

    We produced, selected and cloned hybridomas that secrete monoclonal antibodies against human apolipoprotein (apo) A-I. All of the antibodies corresponded to the IgG(1) subclass and were named 1C11, 2B4, 2C10, 7C5, 8A4 and 8A5. The antibodies were characterized by their reactivity with whole lipoproteins, apolipoproteins, synthetic peptides and fragments generated by cleavage of the apo A-I. Three of the monoclonal antibodies studied (2B4, 2C10 and 7C5) were similarly inhibited by an amino-terminal peptide (amino acid sequence 1-20) of apo A-I, whereas antibodies 1C11, 8A4 and 8A5 had no reaction. Other results show that monoclonal antibody 1C11 recognizes an epitope located between amino acids 135-148. We evaluated the monoclonal antibody 8A4 against different HDL subpopulations by competitive displacement analysis and it showed a similar reactivity with the HDL particles: LpA-I and LpA-I:A-II. This antibody was used to standardize a sandwich ELISA to quantitate LpA-I in plasma. We conclude that these monoclonal antibodies are relevant for the study of apo A-I epitope expression and for quantitating apo A-I containing lipoparticles.

  4. Apolipoprotein A-II-mediated Conformational Changes of Apolipoprotein A-I in Discoidal High Density Lipoproteins*

    PubMed Central

    Gauthamadasa, Kekulawalage; Vaitinadin, Nataraja Sarma; Dressman, James L.; Macha, Stephen; Homan, Reyn; Greis, Kenneth D.; Silva, R. A. Gangani D.

    2012-01-01

    It is well accepted that HDL has the ability to reduce risks for several chronic diseases. To gain insights into the functional properties of HDL, it is critical to understand the HDL structure in detail. To understand interactions between the two major apolipoproteins (apos), apoA-I and apoA-II in HDL, we generated highly defined benchmark discoidal HDL particles. These particles were reconstituted using a physiologically relevant phospholipid, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) incorporating two molecules of apoA-I and one homodimer of apoA-II per particle. We utilized two independent mass spectrometry techniques to study these particles. The techniques are both sensitive to protein conformation and interactions and are namely: 1) hydrogen deuterium exchange combined with mass spectrometry and 2) partial acetylation of lysine residues combined with MS. Comparison of mixed particles with apoA-I only particles of similar diameter revealed that the changes in apoA-I conformation in the presence of apoA-II are confined to apoA-I helices 3–4 and 7–9. We discuss these findings with respect to the relative reactivity of these two particle types toward a major plasma enzyme, lecithin:cholesterol acyltransferase responsible for the HDL maturation process. PMID:22235130

  5. Deep subcortical infarct burden in relation to apolipoprotein B/AI ratio in patients with intracranial atherosclerotic stenosis.

    PubMed

    Park, J-H; Hong, K-S; Lee, J; Kim, Y-J; Song, P

    2013-04-01

    Pre-existing brain infarct (PBI), frequently seen on magnetic resonance imaging and usually silent, is recognized as a risk factor for future stroke. Increased apolipoprotein B (apoB)/apoAI ratio is known to be a risk predictor of ischaemic stroke and is associated with intracranial atherosclerotic stenosis (ICAS). However, little is known about the association of apoB/apoAI ratio with PBI. A total of 522 statin-/fibrate-naïve Korean patients, who experienced acute ischaemic stroke, were categorized into three groups: ICAS (n=254), extracranial (n=51), and no cerebral atherosclerotic stenosis (n=217). We explored the association between apoB/apoAI ratio and PBI lesions according to atherosclerosis type (ICAS, ECAS, and NCAS), PBI location (deep subcortical [ds-PBI] versus hemispheric [h-PBI]), and symptomatic PBI (s-PBI) which was relevant to a prior clinical stroke event. Pre-existing brain infarct(+) patients showed a higher apoB/apoAI ratio than PBI(-) patients (0.81 ± 0.28 vs. 0.72 ± 0.23, P<0.001). In ICAS group, patients with higher apoB/apoAI ratio quartiles had more PBIs, ds-PBIs, and s-PBIs (P=0.020, P=0.025, and P=0.001, respectively). With multivariable analyses, the highest apoB/apoAI ratio quartile was associated with PBI (OR, 2.56; 95% CI, 1.39-4.73), ds-PBI (2.48; 1.33-4.62), and advanced (≥ 3) ds-PBIs (2.68; 1.27-5.63) in ICAS group, but not with h-PBI. s-PBI had a dose-response relationship with apoB/apoAI ratio quartiles (6.18; 1.31-29.13 for the second; 5.34; 1.06-26.83 for the third; and 12.17; 2.50-59.19 for the fourth quartile), when referenced to the first quartile. ApoB/apoAI ratio is associated with asymptomatic deep subcortical ischaemic burden as well as with symptomatic lesion in patients with ICAS. © 2012 The Author(s) European Journal of Neurology © 2012 EFNS.

  6. Plasma lipid transport in the hedgehog: partial characterization of structure and function of apolipoprotein A-I.

    PubMed

    Sparrow, D A; Laplaud, P M; Saboureau, M; Zhou, G; Dolphin, P J; Gotto, A M; Sparrow, J T

    1995-03-01

    Apart from exhibiting the presence of lipoprotein [a] in its plasma, another interest of the European hedgehog in lipoprotein research lies in the quantitative prominence of a complex spectrum of high density lipoproteins (HDL) and very high density lipoproteins (VHDL) as cholesterol transporters in plasma (Laplaud, P. M. et al. 1989. Biochim. Biophys. Acta. 1005: 143-156). We, therefore, initiated studies in the field of reverse cholesterol transport in the hedgehog. As a first step, we characterized apolipoprotein A-I (apoA-I), the main protein component of hedgehog HDL and VHDL. Proteolytic cleavage of apoA-I (M(r) approx. 27 kDa) using two different enzymes resulted in two sets of peptides that were subsequently purified by high performance liquid chromatography, and that allowed us determination of the complete protein sequence. Hedgehog apoA-I thus consists of 241 amino acid residues and exhibits an overall 58% homology to its human counterpart, i.e., the lowest value observed to date among mammalian species. However, it retained the general organization common to all known apoA-Is, i.e., a series of amphipathic helical segments punctuated by proline residues. Circular dichroism experiments indicated a helical content of approx. 45%, increasing to approx. 58% in the presence of lecithin unilamellar liposomes. Apart from other differences, amino acid composition analysis shows that hedgehog apoA-I contains four isoleucine residues, while this amino acid is totally absent from the corresponding protein in higher mammals. Polyclonal antibodies raised against hedgehog apoA-I failed to detect any cross-reactivity between the animal and human proteins, although comparative prediction of the respective antigenic structures using the Hopp-Woods algorithm indicated that several potentially antigenic sites may occur in similar regions of the protein. Finally, hedgehog apoA-I was shown to be able to activate lecithin:cholesterol acyl transferase, although it was 4 to 5

  7. Apolipoprotein A-I Milano exhibits potent antioxidant activity on phospholipid surfaces

    SciTech Connect

    Bielicki, John K.; Oda, Michael N.

    2001-09-21

    Apolipoprotein(apo)A-IMilano and apoA-IParis are rare cysteine variants of apoA-I that produce a HDL deficiency in the absence of cardiovascular disease in humans. This paradox provides the basis for the hypothesis that the cysteine variants posses a beneficial activity not associated with wild-type apoA-I (apoA-IWT). In this study, a unique antioxidant activity of apoA-IMilano and apoA-IParis is described. Antioxidant activity was observed using the monomeric form of the variants and was equally effective before and after initiation of oxidative events. ApoA-IMilano was twice as effective as apoA-IParis in preventing lipoxygenase-mediated oxidation of phospholipids; whereas, apoA-IWT was poorly active. ApoA-IMilano protected phospholipid from reactive oxygen species (ROS) generated via xanthine/xanthine oxidase (X/Xo) but failed to inhibit X/Xo induced reduction of cytochrome C. These results indicate that (1) the antioxidant activity of apoA-IMilano was dependent on phospholipid and (2) the cysteine variant was unable to directly quench ROS in the aqueous phase. There were no differences between lipid-free apoA-IMilano, apoA-IParis, and apoA-IWT in mediating the efflux of cholesterol from macrophages indicating the cysteine variants interacted normally with the ABCA1 efflux pathway. The results indicate that incorporation of a free thiol within an amphipathic alpha helix of apoA-I confers an antioxidant activity distinct from that of apoA-IWT. These studies are the first to relate addition-of-function to rare cysteine mutations in apoA-I primary sequence.

  8. Characterization of High Density Lipoprotein Particles in Familial Apolipoprotein A-I Deficiency With Premature Coronary Atherosclerosis, Corneal Arcus and Opacification, and Tubo-Eruptive and Planar Xanthomas

    USDA-ARS?s Scientific Manuscript database

    We describe two male siblings with homozygous familial apolipoprotein (apo) A-I deficiency, markedly decreased high density lipoprotein (HDL) cholesterol levels, undetectable plasma apoA-1, tubo-eruptive and planar xanthomas, and mild corneal arcus and opacification. Sequencing of the apoA-I gene re...

  9. Apolipoprotein A-I lysine modification: effects on helical content, lipid binding and cholesterol acceptor activity.

    PubMed

    Brubaker, Gregory; Peng, Dao-Quan; Somerlot, Benjamin; Abdollahian, Davood J; Smith, Jonathan D

    2006-01-01

    We examined the role of the positively charged lysine residues in apoAI by chemical modification. Lysine modification by reductive methylation did not alter apoAI's net charge, secondary or tertiary structure as observed by circular dichroism and trytophan fluorescence, respectively, or have much impact on lipid binding or ABCA1-dependent cholesterol acceptor activity. Acetylation of lysine residues lowered the isoelectric point of apoAI, altered its secondary and tertiary structure, and led to a 40% decrease in cholesterol acceptor activity, while maintaining 93% of its lipid binding activity. Exhaustive lysine acetoacetylation lowered apoAI's isoelectric point, profoundly disrupted its secondary and tertiary structure, and led to 90% and 82% reductions in cholesterol acceptor and lipid binding activities, respectively. The dose-dependent acetoacetylation of an increasing proportion of apoAI lysine residues demonstrated that cholesterol acceptor activity was more sensitive to this modification than lipid binding activity, suggesting that apoAI lysine positive charges play an important role in ABCA1 mediated lipid efflux beyond the role needed to maintain alpha-helical content and lipid binding activity.

  10. Concentration of apolipoprotein B is comparable with the apolipoprotein B/apolipoprotein A-I ratio and better than routine clinical lipid measurements in predicting coronary heart disease mortality: findings from a multi-ethnic US population

    PubMed Central

    Sierra-Johnson, Justo; Fisher, Rachel M.; Romero-Corral, Abel; Somers, Virend K.; Lopez-Jimenez, Francisco; Öhrvik, John; Walldius, Göran; Hellenius, Mai-Lis; Hamsten, Anders

    2009-01-01

    Aims Prospective studies indicate that apolipoprotein measurements predict coronary heart disease (CHD) risk; however, evidence is conflicting, especially in the US. Our aim was to assess whether measurements of apolipoprotein B (apoB) and apolipoprotein A-I (apoA-I) can improve the ability to predict CHD death beyond what is possible based on traditional cardiovascular (CV) risk factors and clinical routine lipid measurements. Methods and results We analysed prospectively associations of apolipoprotein measurements, traditional CV risk factors, and clinical routine lipid measurements with CHD mortality in a multi-ethnic representative subset of 7594 US adults (mean age 45 years; 3881 men and 3713 women, median follow-up 124 person-months) from the Third National Health and Nutrition Examination Survey mortality study. Multiple Cox-proportional hazards regression was applied. There were 673 CV deaths of which 432 were from CHD. Concentrations of apoB [hazard ratio (HR) 1.98, 95% confidence interval (CI) 1.09–3.61], apoA-I (HR 0.48, 95% CI 0.27–0.85) and total cholesterol (TC) (HR 1.17, 95% CI 1.02–1.34) were significantly related to CHD death, whereas high density lipoprotein cholesterol (HDL-C) (HR 0.68, 95% CI 0.45–1.05) was borderline significant. Both the apoB/apoA-I ratio (HR 2.14, 95% CI 1.11–4.10) and the TC/HDL-C ratio (HR 1.10, 95% CI 1.04–1.16) were related to CHD death. Only apoB (HR 2.01, 95% CI 1.05–3.86) and the apoB/apoA-I ratio (HR 2.09, 95% CI 1.04–4.19) remained significantly associated with CHD death after adjusting for CV risk factors. Conclusion In the US population, apolipoprotein measurements significantly predict CHD death, independently of conventional lipids and other CV risk factors (smoking, dyslipidaemia, hypertension, obesity, diabetes and C-reactive protein). Furthermore, the predictive ability of apoB alone to detect CHD death was better than any of the routine clinical lipid measurements. Inclusion of apolipoprotein

  11. Safety and Tolerability of CSL112, a Reconstituted, Infusible, Plasma-Derived Apolipoprotein A-I, After Acute Myocardial Infarction

    PubMed Central

    Korjian, Serge; Tricoci, Pierluigi; Daaboul, Yazan; Yee, Megan; Jain, Purva; Alexander, John H.; Steg, P. Gabriel; Lincoff, A. Michael; Kastelein, John J.P.; Mehran, Roxana; D’Andrea, Denise M.; Deckelbaum, Lawrence I.; Merkely, Bela; Zarebinski, Maciej; Ophuis, Ton Oude; Harrington, Robert A.

    2016-01-01

    Background: Human or recombinant apolipoprotein A-I (apoA-I) has been shown to increase high-density lipoprotein–mediated cholesterol efflux capacity and to regress atherosclerotic disease in animal and clinical studies. CSL112 is an infusible, plasma-derived apoA-I that has been studied in normal subjects or those with stable coronary artery disease. This study aimed to characterize the safety, tolerability, pharmacokinetics, and pharmacodynamics of CSL112 in patients with a recent acute myocardial infarction. Methods: The AEGIS-I trial (Apo-I Event Reducing in Ischemic Syndromes I) was a multicenter, randomized, double-blind, placebo-controlled, dose-ranging phase 2b trial. Patients with myocardial infarction were stratified by renal function and randomized 1:1:1 to CSL112 (2 g apoA-I per dose) and high-dose CSL112 (6 g apoA-I per dose), or placebo for 4 consecutive weekly infusions. Coprimary safety end points were occurrence of either a hepatic safety event (an increase in alanine transaminase >3 times the upper limit of normal or an increase in total bilirubin >2 times the upper limit of normal) or a renal safety event (an increase in serum creatinine >1.5 times the baseline value or a new requirement for renal replacement therapy). Results: A total of 1258 patients were randomized, and 91.2% received all 4 infusions. The difference in incidence rates for an increase in alanine transaminase or total bilirubin between both CSL112 arms and placebo was within the protocol-defined noninferiority margin of 4%. Similarly, the difference in incidence rates for an increase in serum creatinine or a new requirement for renal replacement therapy was within the protocol-defined noninferiority margin of 5%. CSL112 was associated with increases in apoA-I and ex vivo cholesterol efflux similar to that achieved in patients with stable coronary artery disease. In regard to the secondary efficacy end point, the risk for the composite of major adverse cardiovascular events

  12. Molecular characterization and developmental expression patterns of apolipoprotein A-I in Senegalese sole (Solea senegalensis Kaup).

    PubMed

    Román-Padilla, J; Rodríguez-Rúa, A; Manchado, M; Hachero-Cruzado, I

    2016-05-01

    The apolipoprotein A-I (ApoA-I) is an essential component of the high density lipoproteins (HDL). In this study, the cDNA and genomic sequences of this apolipoprotein were characterized for first time in Solea senegalensis. The predicted polypeptide revealed conserved structural features including ten repeats in the lipid-binding domain and some residues involved in cholesterol interaction and binding. The gene structure analysis identified four exons and three introns. Moreover, the synteny analysis revealed that apoA-I did not localize with other apolipoproteins indicating a divergent evolution with respect to the apoA-IV and apoE cluster. The phylogenetic analyses identified two distinct apoA-I paralogs in Ostariophysi (referred to as Ia and Ib) and only one (Ib) in Acanthopterygii. Whole-mount in situ hybridization located the apoA-I signal mainly in the yolk syncytial layer in lecitotrophic larval stages. Later at mouth opening, the mRNA signals were detected mainly in liver and intestine compatible with its role in the HDL formation. Moreover, a clear signal was detected in some regions of the brain, retina and neural cord suggesting a role in local regulation of cholesterol homeostasis. After metamorphosis, apoA-I was also detected in other tissues such as gills, head kidney and spleen suggesting a putative role in immunity. Expression analyses in larvae fed two diets with different triacylglycerol levels indicated that apoA-I mRNA levels were more associated to larval size and development than dietary lipid levels. Finally, qPCR analyses of immature and mature transcripts revealed distinct expression profiles suggesting a posttranscriptional regulatory mechanism. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Inhibition of inflammatory signaling pathways in 3T3-L1 adipocytes by apolipoprotein A-I.

    PubMed

    Sultana, Afroza; Cochran, Blake J; Tabet, Fatiha; Patel, Mili; Torres, Luisa Cuesta; Barter, Philip J; Rye, Kerry-Anne

    2016-06-01

    Activation of inflammatory signaling pathways links obesity with metabolic disorders. TLR4-mediated activation of MAPKs and NF-κB are 2 such pathways implicated in obesity-induced inflammation. Apolipoprotein A-I (apoA-I) exerts anti-inflammatory effects on adipocytes by effluxing cholesterol from the cells via the ATP binding cassette transporter A1 (ABCA1). It is not known if these effects involve inhibition of inflammatory signaling pathways by apoA-I. This study asks if apoA-I inhibits activation of MAPKs and NF-κB in mouse 3T3-L1 adipocytes and whether this inhibition is ABCA1 dependent. Incubation of differentiated 3T3-L1 adipocytes with apoA-I decreased cell surface expression of TLR4 by 16 ± 2% and synthesis of the TLR4 adaptor protein, myeloid differentiation primary response 88, by 24 ± 4% in an ABCA1-dependent manner. ApoA-I also inhibited downstream activation of MAPKs, such as ERK, p38MAPK, and JNK, as well as expression of proinflammatory adipokines in bacterial LPS-stimulated 3T3-L1 adipocytes in an ABCA1-dependent manner. ApoA-I, by contrast, suppressed nuclear localization of the p65 subunit of NF-κB by 30 ± 3% in LPS-stimulated 3T3-L1 adipocytes in an ABCA1-independent manner. In conclusion, apoA-I inhibits TLR4-mediated inflammatory signaling pathways in adipocytes by preventing MAPK and NF-κB activation.-Sultana, A., Cochran, B. J., Tabet, F., Patel, M., Cuesta Torres, L., Barter, P. J., Rye, K.-A. Inhibition of inflammatory signaling pathways in 3T3-L1 adipocytes by apolipoprotein A-I. © FASEB.

  14. Structures of apolipoprotein A-I in high density lipoprotein generated by electron microscopy and biased simulations.

    PubMed

    Zhu, Lin; Petrlova, Jitka; Gysbers, Peter; Hebert, Hans; Wallin, Stefan; Jegerschöld, Caroline; Lagerstedt, Jens O

    2017-07-25

    Apolipoprotein A-I (apoA-I) in high-density lipoprotein (HDL) is a key protein for the transport of cholesterol from the vascular wall to the liver. The formation and structure of nascent HDL, composed of apoA-I and phospholipids, is critical to this process. The HDL was assembled in vitro from apoA-I, cholesterol and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) at a 1:4:50 molar ratio. The structure of HDL was investigated in vitreous samples, frozen at cryogenic temperatures, as well as in negatively stained samples by transmission electron microscopy. Low resolution electron density maps were next used as restraints in biased Monte Carlo simulations of apolipoprotein A-I dimers, with an initial structure derived from atomic resolution X-ray structures. Two final apoA-I structure models for the full-length structure of apoA-I dimer in the lipid bound conformation were generated, showing a nearly circular, flat particle with an uneven particle thickness. The generated structures provide evidence for the discoidal, antiparallel arrangement of apoA-I in nascent HDL, and propose two preferred conformations of the flexible N-termini. The novel full-length structures of apoA-I dimers deepens the understanding to the structure-function relationship of nascent HDL with significance for the prevention of lipoprotein-related disease. The biased simulation method used in this study provides a powerful and convenient modelling tool with applicability for structural studies and modelling of other proteins and protein complexes. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Homocysteine diminishes apolipoprotein A-I function and expression in patients with hypothyroidism: a cross-sectional study.

    PubMed

    Yang, Ning; Yao, Zhi; Miao, Li; Liu, Jia; Gao, Xia; Xu, Yuan; Wang, Guang

    2016-07-26

    Hypothyroidism (HO) can significantly impair lipid metabolism and increase cardiovascular disease risk. Hyperhomocysteinemia (HHcy) is an independent risk factor for cardiovascular disease. Our previous study demonstrated that HHcy significantly induced insulin resistance and impaired coronary artery endothelial function in patients with either hypertension or HO. In the present study, we studied whether plasma levels of high-density lipoprotein-cholesterol (HDL-C) and apolipoprotein A-I (Apo A-I) were altered in patients with HO, and if so, whether this change was mediated by HHcy. A total of 258 subjects were enrolled and divided into the following three groups: control group (n = 94), HO group (n = 73), and subclinical hypothyroidism (SHO) group (n = 91). Additionally, all groups were subdivided based on the subjects' Hcy levels into HHcy (plasma Hcy level over 15 μmol/l) and normal Hcy subgroups. The plasma levels of lipid indexes were measured. Statistical analyses were performed to evaluate the correlations between groups. The plasma Hcy levels were significantly higher in the HO group than in the SHO or control groups (all p < 0.05). Moreover, levels of Apo A-I and HDL-C were markedly reduced in the HHcy subgroup compared with the normal Hcy subgroup for patients with either HO (Apo A-I: p < 0.05; HDL-C: p < 0.01) or SHO (Apo A-I: p < 0.05; HDL-C: p < 0.01). In addition, the plasma Hcy levels were negatively correlated with levels of Apo A-I in all three groups (HO group: r = - 0.320, SHO group: r = - 0.337 and control group: r = - 0.317; all p < 0.01). Hcy levels were significantly increased in patients with HO or SHO. These increased Hcy levels may impair cardiovascular function via the inhibition of Apo A-1 expression and impairment of its antioxidant capacity. Our findings provide new insights into the pathogenesis of hypothyroidism-induced metabolic disorders.

  16. Human Frontal Lobes and AI Planning Systems

    NASA Technical Reports Server (NTRS)

    Levinson, Richard; Lum, Henry Jr. (Technical Monitor)

    1994-01-01

    Human frontal lobes are essential for maintaining a self-regulating balance between predictive and reactive behavior. This paper describes a system that integrates prediction and reaction based on neuropsychological theories of frontal lobe function. In addition to enhancing our understanding of deliberate action in humans' the model is being used to develop and evaluate the same properties in machines. First, the paper presents some background neuropsychology in order to set a general context. The role of frontal lobes is then presented by summarizing three theories which formed the basis for this work. The components of an artificial frontal lobe are then discussed from both neuropsychological and AI perspectives. The paper concludes by discussing issues and methods for evaluating systems that integrate planning and reaction.

  17. Human Frontal Lobes and AI Planning Systems

    NASA Technical Reports Server (NTRS)

    Levinson, Richard; Lum, Henry Jr. (Technical Monitor)

    1994-01-01

    Human frontal lobes are essential for maintaining a self-regulating balance between predictive and reactive behavior. This paper describes a system that integrates prediction and reaction based on neuropsychological theories of frontal lobe function. In addition to enhancing our understanding of deliberate action in humans' the model is being used to develop and evaluate the same properties in machines. First, the paper presents some background neuropsychology in order to set a general context. The role of frontal lobes is then presented by summarizing three theories which formed the basis for this work. The components of an artificial frontal lobe are then discussed from both neuropsychological and AI perspectives. The paper concludes by discussing issues and methods for evaluating systems that integrate planning and reaction.

  18. Chromatofocusing of apolipoproteins from human serum high density lipoprotein.

    PubMed

    Knipping, G; Steyrer, E; Holasek, A

    1984-01-01

    Human HDL was delipidated and the apolipoproteins were fractionated by chromatofocusing. Chromatofocusing, which separates proteins due to their differing isoelectric points, resulted in 8 peaks with corresponding pI values of 7.40, 6.92, 6.64, 5.48, 5.30, 5.18, 4.92 and 4.63. By one single chromatofocusing run four apolipoproteins were obtained in pure form. Two additional polypeptides could be purified during the desalting step using phenyl-Sepharose.

  19. Screening and identification of apolipoprotein A-I as a potential hepatoblastoma biomarker in children, excluding inflammatory factors.

    PubMed

    Zhao, Wei; Li, Juan; Zhang, Yilin; Gao, Pengfei; Zhang, Junjie; Guo, Fei; Yu, Jiekai; Zheng, Shu; Wang, Jiaxiang

    2015-07-01

    The aim of the present study was to identify a child hepatoblastoma serum biomarker that is unaffected by inflammatory factors, with the ultimate aim of finding an effective method for the early diagnosis of hepatoblastoma. The magnetic bead-based weak cation exchange chromatography technique was used to process serum harvested from 30 children with hepatoblastoma, 20 children with systemic inflammatory response syndrome (SIRS) and 20 healthy children. Proteins differentially expressed in SIRS were excluded from consideration as biomarkers for hepatoblastoma. Proteins differentially expressed in hepatoblastoma and healthy controls were screened using surface-enhanced laser desorption/ionization-time of flight-mass spectrometry (SELDI-TOF-MS). Target proteins were purified by SDS-PAGE, and matrix-assisted laser desorption/ionization (MALDI)-TOF-MS was used to determine their amino acid sequences. Protein matches were searched in the SwissProt database. Quantitative polymerase chain reaction (qPCR) and ELISA were employed to confirm the expression of target proteins. Following screening to exclude inflammatory factors, SELDI-TOF-MS revealed a protein with a mass-to-charge ratio of 9,348 Da that was expressed at significantly lower levels in the serum of children with hepatoblastoma compared with healthy controls (P<0.01). Sequence analysis identified this protein as apolipoprotein A-1 (Apo A-I). qPCR and ELISA confirmed that the expression of Apo A-I mRNA and protein were significantly lower in children with hepatoblastoma compared with healthy controls (P<0.05). These results indicate that Apo A-I is a non-inflammatory protein marker for hepatoblastoma with the potential for use in early diagnosis of hepatoblastoma. In addition, the present study demonstrates the feasibility of proteomic screening for the identification of proteins that can serve as markers for a specific tumor.

  20. Screening and identification of apolipoprotein A-I as a potential hepatoblastoma biomarker in children, excluding inflammatory factors

    PubMed Central

    ZHAO, WEI; LI, JUAN; ZHANG, YILIN; GAO, PENGFEI; ZHANG, JUNJIE; GUO, FEI; YU, JIEKAI; ZHENG, SHU; WANG, JIAXIANG

    2015-01-01

    The aim of the present study was to identify a child hepatoblastoma serum biomarker that is unaffected by inflammatory factors, with the ultimate aim of finding an effective method for the early diagnosis of hepatoblastoma. The magnetic bead-based weak cation exchange chromatography technique was used to process serum harvested from 30 children with hepatoblastoma, 20 children with systemic inflammatory response syndrome (SIRS) and 20 healthy children. Proteins differentially expressed in SIRS were excluded from consideration as biomarkers for hepatoblastoma. Proteins differentially expressed in hepatoblastoma and healthy controls were screened using surface-enhanced laser desorption/ionization-time of flight-mass spectrometry (SELDI-TOF-MS). Target proteins were purified by SDS-PAGE, and matrix-assisted laser desorption/ionization (MALDI)-TOF-MS was used to determine their amino acid sequences. Protein matches were searched in the SwissProt database. Quantitative polymerase chain reaction (qPCR) and ELISA were employed to confirm the expression of target proteins. Following screening to exclude inflammatory factors, SELDI-TOF-MS revealed a protein with a mass-to-charge ratio of 9,348 Da that was expressed at significantly lower levels in the serum of children with hepatoblastoma compared with healthy controls (P<0.01). Sequence analysis identified this protein as apolipoprotein A-1 (Apo A-I). qPCR and ELISA confirmed that the expression of Apo A-I mRNA and protein were significantly lower in children with hepatoblastoma compared with healthy controls (P<0.05). These results indicate that Apo A-I is a non-inflammatory protein marker for hepatoblastoma with the potential for use in early diagnosis of hepatoblastoma. In addition, the present study demonstrates the feasibility of proteomic screening for the identification of proteins that can serve as markers for a specific tumor. PMID:26171005

  1. Association of serum lipids and coronary artery disease with polymorphisms in the apolipoprotein AI-CIII-AIV gene cluster.

    PubMed

    Rai, Himanshu; Sinha, Nakul; Finn, James; Agrawal, Suraksha; Mastana, Sarabjit

    2016-12-31

    Genetic variants are considered as one of the main determinants of the concentration of serum lipids and coronary artery disease (CAD). Polymorphisms in the Apolipoprotein (Apo) AI-CIII-AIV gene cluster has been known to affect the concentrations of various lipid sub-fractions and the risk of CAD. The present study assessed associations between polymorphisms of the Apo AI-CIII-AIV gene cluster, [ApoA-I,-75G > A, (rs1799837); ApoC-III 3238C > G, (SstI), (rs5128) and ApoA-IV, Thr347Ser(347A > T), (rs675)] with serum lipids and their contributions to CAD in North Indian population. We recruited age, sex matched, 200 CAD patients and 200 healthy controls and tested them for fasting levels of serum lipids. We genotyped selected polymorphisms using polymerase chain reaction-restriction fragment length polymorphism. There were no statistically significant association of selected polymorphisms (or their combinations) with CAD even after employing additive, dominant and recessive models. However there was significant association of selected polymorphisms with various lipid traits amongst the control cohort (p < 0.05). Mean levels of high density lipoprotein cholesterol and triglycerides were found to be significantly higher among controls carrying at least one mutant allele at ApoA1-75G > A (p = 0.019) and ApoCIII SstI (p < 0.001) polymorphism respectively. Our study observed that the selected polymorphisms in the ApoAI-CIII-AIV gene cluster although significantly affect various lipid traits but this affect does not seem to translate into association with CAD, at least among North Indian population.

  2. Association of serum lipids and coronary artery disease with polymorphisms in the apolipoprotein AI-CIII-AIV gene cluster

    PubMed Central

    Rai, Himanshu; Sinha, Nakul; Finn, James; Agrawal, Suraksha; Mastana, Sarabjit

    2016-01-01

    Abstract Genetic variants are considered as one of the main determinants of the concentration of serum lipids and coronary artery disease (CAD). Polymorphisms in the Apolipoprotein (Apo) AI-CIII-AIV gene cluster has been known to affect the concentrations of various lipid sub-fractions and the risk of CAD. The present study assessed associations between polymorphisms of the Apo AI-CIII-AIV gene cluster, [ApoA-I,-75G > A, (rs1799837); ApoC-III 3238C > G, (SstI), (rs5128) and ApoA-IV, Thr347Ser(347A > T), (rs675)] with serum lipids and their contributions to CAD in North Indian population. We recruited age, sex matched, 200 CAD patients and 200 healthy controls and tested them for fasting levels of serum lipids. We genotyped selected polymorphisms using polymerase chain reaction-restriction fragment length polymorphism. There were no statistically significant association of selected polymorphisms (or their combinations) with CAD even after employing additive, dominant and recessive models. However there was significant association of selected polymorphisms with various lipid traits amongst the control cohort (p < 0.05). Mean levels of high density lipoprotein cholesterol and triglycerides were found to be significantly higher among controls carrying at least one mutant allele at ApoA1-75G > A (p = 0.019) and ApoCIII SstI (p < 0.001) polymorphism respectively. Our study observed that the selected polymorphisms in the ApoAI-CIII-AIV gene cluster although significantly affect various lipid traits but this affect does not seem to translate into association with CAD, at least among North Indian population. PMID:28261635

  3. Associations of apolipoprotein B/apolipoprotein A-I ratio with pre-diabetes and diabetes risks: a cross-sectional study in Chinese adults.

    PubMed

    Zheng, Shuang; Han, Tingting; Xu, Hua; Zhou, Huan; Ren, Xingxing; Wu, Peihong; Zheng, Jun; Wang, Lihua; Zhang, Ming; Jiang, Yihong; Chen, Yawen; Qiu, Huiying; Liu, Wei; Hu, Yaomin

    2017-01-20

    Apolipoprotein B/apolipoprotein A-I (ApoB/ApoA-I) ratio is a useful predictor of cardiovascular risk. However, the association between the ApoB/ApoA-I ratio and the risk of type 2 diabetes mellitus (T2DM) is still obscure. To investigate the associations between the ApoB/ApoA-I ratio and the risk of T2DM and pre-diabetes in a Chinese population, and to assess the role of gender in these associations. A stratified random sampling design was used in this cross-sectional study which included 264 men and 465 women with normal glucose tolerance (NGT), pre-diabetes or T2DM. Serum ApoB, ApoA-I and other lipid and glycaemic traits were measured. Pearson's partial correlation and multivariable logistic analysis were used to evaluate the associations between ApoB/ApoA-I ratio and the risk of T2DM and pre-diabetes. The ApoB/ApoA-I ratios were significantly increased across the spectrum of NGT, pre-diabetes and T2DM. Women showed higher levels of ApoB/ApoA-I ratio and ApoB than men in the pre-diabetic and T2DM groups, but not in the NGT group. The ApoB/ApoA-I ratio was closely related with triglyceride, total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and other glycaemic traits. Moreover, in women, the risk of diabetes and pre-diabetes in the top and middle tertiles of the ApoB/ApoA-I ratio were 3.65-fold (95% CI 1.69 to 6.10) and 2.19-fold (95% CI 1.38 to 2.84) higher than in the bottom tertile, respectively, after adjusting for potential confounding factors. However, the associations disappeared in men after adjusting for other factors. The ApoB/ApoA-I ratio showed positive associations with the risk of diabetes and pre-diabetes in Chinese women. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  4. Associations of apolipoprotein B/apolipoprotein A-I ratio with pre-diabetes and diabetes risks: a cross-sectional study in Chinese adults

    PubMed Central

    Zheng, Shuang; Han, Tingting; Xu, Hua; Zhou, Huan; Ren, Xingxing; Wu, Peihong; Zheng, Jun; Wang, Lihua; Zhang, Ming; Jiang, Yihong; Chen, Yawen; Qiu, Huiying; Liu, Wei; Hu, Yaomin

    2017-01-01

    Background Apolipoprotein B/apolipoprotein A-I (ApoB/ApoA-I) ratio is a useful predictor of cardiovascular risk. However, the association between the ApoB/ApoA-I ratio and the risk of type 2 diabetes mellitus (T2DM) is still obscure. Aims To investigate the associations between the ApoB/ApoA-I ratio and the risk of T2DM and pre-diabetes in a Chinese population, and to assess the role of gender in these associations. Methods A stratified random sampling design was used in this cross-sectional study which included 264 men and 465 women with normal glucose tolerance (NGT), pre-diabetes or T2DM. Serum ApoB, ApoA-I and other lipid and glycaemic traits were measured. Pearson's partial correlation and multivariable logistic analysis were used to evaluate the associations between ApoB/ApoA-I ratio and the risk of T2DM and pre-diabetes. Results The ApoB/ApoA-I ratios were significantly increased across the spectrum of NGT, pre-diabetes and T2DM. Women showed higher levels of ApoB/ApoA-I ratio and ApoB than men in the pre-diabetic and T2DM groups, but not in the NGT group. The ApoB/ApoA-I ratio was closely related with triglyceride, total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol and other glycaemic traits. Moreover, in women, the risk of diabetes and pre-diabetes in the top and middle tertiles of the ApoB/ApoA-I ratio were 3.65-fold (95% CI 1.69 to 6.10) and 2.19-fold (95% CI 1.38 to 2.84) higher than in the bottom tertile, respectively, after adjusting for potential confounding factors. However, the associations disappeared in men after adjusting for other factors. Conclusions The ApoB/ApoA-I ratio showed positive associations with the risk of diabetes and pre-diabetes in Chinese women. PMID:28110289

  5. Macrophage apolipoprotein A-I expression protects against atherosclerosis in ApoE-deficient mice and up-regulates ABC transporters.

    PubMed

    Su, Yan Ru; Ishiguro, Hiroyuki; Major, Amy S; Dove, Dwayne E; Zhang, Wenwu; Hasty, Alyssa H; Babaev, Vladimir R; Linton, MacRae F; Fazio, Sergio

    2003-10-01

    The antiatherogenic effect of high-density lipoprotein (HDL) and its major protein component apolipoprotein A-I (apoA-I) has been largely attributed to their key roles in reverse cholesterol transport (RCT) and cellular cholesterol efflux. Substantial evidence shows that overexpression of human apoA-I reduces atherosclerosis in animal models. However, it is uncertain whether this protection is due to an increase in plasma HDL level or to a local effect in the artery wall. To test the hypothesis that expression of human apoA-I in macrophages can promote RCT in the artery wall, we used a retroviral construct expressing human apoA-I cDNA (MFG-HAI) to transduce ApoE(-/-) bone marrow cells and then transplanted these cells into ApoE(-/-) mice with preexisting atherosclerosis. ApoE(-/-) mice reconstituted with MFG-HAI marrow had a significant reduction (30%) in atherosclerotic lesions in the proximal aorta compared to control mice that received marrow expressing MFG parental virus. Peritoneal macrophages isolated from MFG-HAI mice showed a four- to fivefold increase in mRNA expression levels of both ATP-binding cassette (ABC) A1 and ABCG1 compared to controls. Our data demonstrate that gene transfer-mediated expression of human apoA-I in macrophages can compensate in part for apoE deficiency and delay the progression of atherosclerotic lesions by stimulating ABC-dependent cholesterol efflux and RCT.

  6. Apolipoprotein A-I Mimetic Peptides: Discordance Between In Vitro and In Vivo Properties-Brief Report.

    PubMed

    Ditiatkovski, Michael; Palsson, Jonatan; Chin-Dusting, Jaye; Remaley, Alan T; Sviridov, Dmitri

    2017-07-01

    Apolipoprotein A-I (apoA-I) mimetic peptides have antiatherogenic properties of high-density lipoprotein in vitro and have been shown to inhibit atherosclerosis in vivo. It is unclear, however, if each in vitro antiatherogenic property of these peptides translates to a corresponding activity in vivo, and if so, which of these contributes most to reduce atherosclerosis. The effect of 7 apoA-I mimetic peptides, which were developed to selectively reproduce a specific component of the antiatherogenic properties of apoA-I, on the development of atherosclerosis was investigated in apolipoprotein E-deficient mice fed a high-fat diet for 4 or 12 weeks. The peptides include those that selectively upregulate cholesterol efflux, or are anti-inflammatory, or have antioxidation properties. All the peptides studied effectively inhibited the in vivo development of atherosclerosis in this model to the same extent. However, none of the peptides had the same selective effect in vivo as they had exhibited in vitro. None of the tested peptides affected plasma lipoprotein profile; capacity of plasma to support cholesterol efflux was increased modestly and similarly for all peptides. There is a discordance between the selective in vitro and in vivo functional properties of apoA-I mimetic peptides, and the in vivo antiatherosclerotic effect of apoA-I-mimetic peptides is independent of their in vitro functional profile. Comparing the properties of apoA-I mimetic peptides in plasma rather than in the lipid-free state is better for predicting their in vivo effects on atherosclerosis. © 2017 American Heart Association, Inc.

  7. Elevated high density lipoprotein cholesterol levels correlate with decreased apolipoprotein A-I and A-II fractional catabolic rate in women.

    PubMed Central

    Brinton, E A; Eisenberg, S; Breslow, J L

    1989-01-01

    High levels of HDL-cholesterol (HDL-C) protect against coronary heart disease susceptibility, but the metabolic mechanisms underlying elevated HDL-C levels are poorly understood. We now report the turnover of isologous radioiodinated HDL apolipoproteins, apo A-I and apo A-II, in 15 female subjects on a metabolic diet with HDL-C levels ranging from 51 to 122 mg/dl. The metabolic parameters, fractional catabolic rate (FCR) and absolute synthetic rate (SR), were determined for apo A-I and apo A-II in all subjects. There was an inverse correlation between plasma HDL-C and the FCR of apo A-I and apo A-II (r = -0.75, P less than 0.001, and r = -0.54, P = 0.036, respectively), but no correlation with the SR of either apo A-I or apo A-II (r = 0.09, and r = -0.16, respectively, both P = NS). Apo A-I levels correlated inversely with apo A-I FCR (r = -0.64, P = 0.01) but not with apo A-I SR (r = 0.30, P = NS). In contrast, plasma levels of apo A-II did not correlate with apo A-II FCR (r = -0.38, P = 0.16), but did correlate with apo A-II SR (r = 0.65, P = 0.009). Further analysis showed that apo A-I and apo A-II FCR were inversely correlated with the HDL-C/apo A-I + A-II ratio (r = -0.69 and -0.61, P = 0.005 and 0.015, respectively). These data suggest that: (a) low HDL apolipoprotein FCR is the predominant metabolic mechanism of elevated HDL-C levels; (b) apo A-I FCR is the primary factor in controlling plasma apo A-I levels, but apo A-II SR is the primary factor controlling plasma apo A-II levels; (c) low HDL apolipoprotein FCR is associated with a lipid-rich HDL fraction. These findings elucidate aspects of HDL metabolism which contribute to high HDL-C levels and which may constitute mechanisms for protection against coronary heart disease. PMID:2500457

  8. High-density cholesterol and apolipoprotein AI as modifiers of plasma fibrin clot properties in apparently healthy individuals.

    PubMed

    Ząbczyk, Michał; Hońdo, Łukasz; Krzek, Marzena; Undas, Anetta

    2013-01-01

    Low high-density lipoprotein cholesterol (HDL-C) increases cardiovascular risk, whereas its high levels protect against atherosclerosis via multiple beneficial effects. Dense and poorly lysable fibrin clot formation is observed in cardiovascular disease. We sought to investigate whether HDL-C and its major component apolipoprotein A (Apo A)-I affect fibrin clot properties. In 136 apparently healthy individuals (99 men, 37 women, aged 49-69 years) we determined plasma fibrin clot permeability (Ks coefficient) and lysis time (t50%) together with Apo A-I and lipoprotein (a) [Lp(a)] levels. The median HDL-C level was 1.33  mmol/l (range from 0.77 to 2.19  mmol/l). HDL-C was positively associated with Apo A-I (r = 0.62, P < 0.00001). HDL-C and Apo A-I were positively correlated with Ks (r = 0.52, P < 0.00001 and r = 0.44, P < 0.00001, respectively) and inversely with t50% (r = -0.44, P < 0.00001 and r = -0.35, P = 0.00003, respectively). No such associations were seen for other lipid variables. Ks and t50% were associated with Lp(a) (r = -0.42, P < 0.00001 and r = 0.42, P < 0.00001, respectively) and fibrinogen (r = -0.31, P = 0.00024 and r = 0.39, P < 0.00001, respectively). Individuals with HDL-C at least 1.4 mmol/l (n = 54) had 19% higher Ks (P = 0.00016) and 17% shorter t50% (P = 0.0012) than the remainder. After adjustment for age, fibrinogen, and Lp(a), HDL-C was the independent predictor of Ks (β = 0.7, P < 0.00001) and t50% (β = -0.62, P < 0.00001). This study shows that elevated HDL-C levels are associated with improved fibrin clot permeability and lysis, indicating a novel antithrombotic mechanism underlying the postulated beneficial effects of therapy targeted at HDL-C.

  9. A mass spectrometric determination of the conformation of dimeric apolipoprotein A-I in discoidal high density lipoproteins.

    PubMed

    Silva, R A Gangani D; Hilliard, George M; Li, Ling; Segrest, Jere P; Davidson, W Sean

    2005-06-21

    Discoidal forms of high density lipoproteins (HDL) are critical intermediates between lipid-poor apolipoprotein A-I (apo A-I), the major protein constituent of HDL, and the mature spherical forms that comprise the bulk of circulating particles. Thus, many studies have focused on understanding apoA-I structure in discs reconstituted in vitro. Recent theoretical and experimental work supports a "belt" model for apoA-I in which repeating amphipathic helical domains run parallel to the plane of the lipid disc. However, disc-associated apoA-I can adopt several tertiary arrangements that are consistent with a belt orientation. To distinguish among these, we cross-linked near-neighbor Lys groups in homogeneous 96 A discs containing exactly two molecules of apoA-I. After delipidation and tryptic digestion, mass spectrometry was used to identify 9 intermolecular and 11 intramolecular cross-links. The cross-linking pattern strongly suggests a "double-belt" molecular arrangement for apoA-I in which two apoA-I molecules wrap around the lipid bilayer disc forming two stacked rings in an antiparallel orientation with helix 5 of each apoA-I in juxtaposition (LL5/5 orientation). The data also suggests the presence of an additional double-belt orientation with a shifted helical registry (LL5/2 orientation). Furthermore, a 78 A particle with two molecules of apoA-I fit a similar double-belt motif with evidence for conformational changes in the N-terminus and the region near helix 5. A comparison of this work to a previous study is suggestive that a third molecule of apoA-I can form a hairpin in larger particles containing three molecules of apoA-I.

  10. High-density Lipoproteins and Apolipoprotein A-I: Potential New Players in the Prevention and Treatment of Lung Disease.

    PubMed

    Gordon, Elizabeth M; Figueroa, Debbie M; Barochia, Amisha V; Yao, Xianglan; Levine, Stewart J

    2016-01-01

    Apolipoprotein A-I (apoA-I) and high-density lipoproteins (HDL) mediate reverse cholesterol transport out of cells. Furthermore, HDL has additional protective functions, which include anti-oxidative, anti-inflammatory, anti-apoptotic, and vasoprotective effects. In contrast, HDL can become dysfunctional with a reduction in both cholesterol efflux and anti-inflammatory properties in the setting of disease or the acute phase response. These paradigms are increasingly being recognized to be active in the pulmonary system, where apoA-I and HDL have protective effects in normal lung health, as well as in a variety of disease states, including acute lung injury (ALI), asthma, chronic obstructive pulmonary disease, lung cancer, pulmonary arterial hypertension, pulmonary fibrosis, and viral pneumonia. Similar to observations in cardiovascular disease, however, HDL may become dysfunctional and contribute to disease pathogenesis in respiratory disorders. Furthermore, synthetic apoA-I mimetic peptides have been shown to have protective effects in animal models of ALI, asthma, pulmonary hypertension, and influenza pneumonia. These findings provide evidence to support the concept that apoA-I mimetic peptides might be developed into a new treatment that can either prevent or attenuate the manifestations of lung diseases, such as asthma. Thus, the lung is positioned to take a page from the cardiovascular disease playbook and utilize the protective properties of HDL and apoA-I as a novel therapeutic approach.

  11. Decreased apolipoprotein A-I level indicates poor prognosis in extranodal natural killer/T-cell lymphoma, nasal type.

    PubMed

    Quan, Qi; Chen, Qi; Chen, Ping; Jiang, Li; Li, Tingwei; Qiu, Huijuan; Zhang, Bei

    2016-01-01

    Extranodal natural killer (NK)/T-cell lymphoma, nasal type (ENKTL) is an invasive lymphoid malignancy with unfavorable survival, for which a prognostic model has not yet been validated. We hypothesized that serum apolipoprotein A-I (ApoA-I) may serve as a novel prognostic marker for ENKTL. A total of 236 newly diagnosed cases of ENKTL were analyzed retrospectively. The optimal cutoff value for the serum ApoA-I level was determined to be 0.95 g/L. A total of 154 and 82 cases were assigned to the high and low ApoA-I groups, respectively. Patients in the low ApoA-I group tended to present with poorer clinical features, a lower complete remission rate (P=0.001), and poor median progression-free survival (P<0.001) and overall survival (P<0.001). Multivariate analysis using Cox model showed that the serum ApoA-I level was an independent prognostic marker of overall survival (P<0.001) and progression-free survival (P<0.001) for ENKTL patients. For cases in the low-risk group, as assessed by International Prognostic Index, Prognosis Index for peripheral T-cell lymphoma, unspecified, and Korean Prognostic Index, the serum ApoA-I level was able to differentiate cases with poor outcomes from cases with good outcomes. Our results showed that the baseline serum ApoA-I level was helpful for predicting ENKTL prognosis.

  12. Plasma activated coating immobilizes apolipoprotein A-I to stainless steel surfaces in its bioactive form and enhances biocompatibility.

    PubMed

    Vanags, Laura Z; Tan, Joanne T M; Santos, Miguel; Michael, Praveesuda S; Ali, Ziad; Bilek, Marcela M M; Wise, Steven G; Bursill, Christina A

    2017-06-29

    We utilized a plasma activated coating (PAC) to covalently bind the active component of high density lipoproteins (HDL), apolipoprotein (apo) A-I, to stainless steel (SS) surfaces. ApoA-I suppresses restenosis and thrombosis and may therefore improve SS stent biocompatibility. PAC-coated SS significantly increased the covalent attachment of apoA-I, compared to SS alone. In static and dynamic flow thrombosis assays, PAC+apoA-I inhibited thrombosis and reduced platelet activation marker p-selectin. PAC+apoA-I reduced smooth muscle cell attachment and proliferation, and augmented EC attachment to PAC. We then coated PAC onto murine SS stents and found it did not peel or delaminate following crimping/expansion. ApoA-I was immobilized onto PAC-SS stents and was retained as a monolayer when exposed to pulsatile flow in vivo in a murine stent model. In conclusion, ApoA-I immobilized on PAC withstands pulsatile flow in vivo and retains its bioactivity, exhibiting anti-thrombotic and anti-restenotic properties, demonstrating the potential to improve stent biocompatibility. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Seasonal variation in plasma lipids, lipoproteins, apolipoprotein A-I and vitellogenin in the freshwater turtle, Chrysemys picta.

    PubMed

    Duggan, A; Paolucci, M; Tercyak, A; Gigliotti, M; Small, D; Callard, I

    2001-09-01

    An analysis of plasma lipids and lipoprotein fractions was performed over the course of the annual ovarian cycle of the female turtle, Chrysemys picta. Determinations of total plasma triglycerides, cholesterol, vitellogenin and apolipoprotein A-I (apoA-I) were made. The lipid and protein composition of the lipoprotein fractions [very low density lipoprotein (VLDL), low density lipoprotein (LDL), high density lipoprotein (HDL) and very high density lipoprotein (VHDL)] were also observed over the same period. Plasma triglyceride and vitellogenin levels were significantly increased in the spring preovulatory period and fall recrudescent phase. Total plasma cholesterol levels were significantly elevated only at the onset of the fall recrudescent phase and apoA-I levels were highest during the postoviposition/ovarian arrest phase. The triglyceride content of VLDL was highest in preovulatory animals and there were apparent seasonal changes in the expression of apoA-I and apoE of HDL/VHDL. We conclude that the coordinate regulation of lipids and protein contributes to seasonal ovarian growth and clearance of lipids from plasma, both of which are most likely under hormonal control.

  14. High-density Lipoproteins and Apolipoprotein A-I: Potential New Players in the Prevention and Treatment of Lung Disease

    PubMed Central

    Gordon, Elizabeth M.; Figueroa, Debbie M.; Barochia, Amisha V.; Yao, Xianglan; Levine, Stewart J.

    2016-01-01

    Apolipoprotein A-I (apoA-I) and high-density lipoproteins (HDL) mediate reverse cholesterol transport out of cells. Furthermore, HDL has additional protective functions, which include anti-oxidative, anti-inflammatory, anti-apoptotic, and vasoprotective effects. In contrast, HDL can become dysfunctional with a reduction in both cholesterol efflux and anti-inflammatory properties in the setting of disease or the acute phase response. These paradigms are increasingly being recognized to be active in the pulmonary system, where apoA-I and HDL have protective effects in normal lung health, as well as in a variety of disease states, including acute lung injury (ALI), asthma, chronic obstructive pulmonary disease, lung cancer, pulmonary arterial hypertension, pulmonary fibrosis, and viral pneumonia. Similar to observations in cardiovascular disease, however, HDL may become dysfunctional and contribute to disease pathogenesis in respiratory disorders. Furthermore, synthetic apoA-I mimetic peptides have been shown to have protective effects in animal models of ALI, asthma, pulmonary hypertension, and influenza pneumonia. These findings provide evidence to support the concept that apoA-I mimetic peptides might be developed into a new treatment that can either prevent or attenuate the manifestations of lung diseases, such as asthma. Thus, the lung is positioned to take a page from the cardiovascular disease playbook and utilize the protective properties of HDL and apoA-I as a novel therapeutic approach. PMID:27708582

  15. Effect of lipid-bound apolipoprotein A-I cysteine mutant on ATF3 in RAW264.7 cells

    PubMed Central

    Wang, Yunlong; Wang, Yanhui; Jia, Shaoyou; Dong, Qingzhe; Chen, Yuanbin

    2017-01-01

    Activating transcription factor 3 (ATF3) is a TLR-induced repressor that plays an important role in the inhibition of specific inflammatory signals. We previously constructed recombinant high density lipoproteins (rHDL) (including rHDLWT, rHDLM, rHDL228 and rHDL74) and found that rHDL74 had a strong anti-inflammatory ability. In the present study, we investigate the roles of recombinant apolipoprotein A-I (ApoA-I) (rHDLWT) and its cysteine mutant HDLs (rHDLM, rHDL228 and rHDL74) on ATF3 function in RAW264.7 cells stimulated by lipopolysaccharide. Our results showed that compared with the LPS group, rHDL74 can decrease the level of TNF-α and IL-6, whereas rHDL228 increases their expression levels. RT-PCR and Western blotting results showed that compared with the LPS group, rHDL74, rHDLWT and rHDLM can markedly increase the expression level of ATF3, whereas the level of ATF3 decreases in the rHDL228 group. In summary, the different anti-inflammatory mechanisms of the ApoA-I cysteine mutants might be associated with the regulation of ATF3 level. PMID:28093456

  16. Interaction of thioflavin T with amyloid fibrils of apolipoprotein A-I N-terminal fragment: resonance energy transfer study.

    PubMed

    Girych, Mykhailo; Gorbenko, Galyna; Trusova, Valeriya; Adachi, Emi; Mizuguchi, Chiharu; Nagao, Kohjiro; Kawashima, Hiroyuki; Akaji, Kenichi; Lund-Katz, Sissel; Phillips, Michael C; Saito, Hiroyuki

    2014-01-01

    Apolipoprotein A-I is amenable to a number of specific mutations associated with hereditary systemic amyloidoses. Amyloidogenic properties of apoA-I are determined mainly by its N-terminal fragment. In the present study Förster resonance energy transfer between tryptophan as a donor and Thioflavin T as an acceptor was employed to obtain structural information on the amyloid fibrils formed by apoA-I variant 1-83/G26R/W@8. Analysis of the dye-fibril binding data provided evidence for the presence of two types of ThT binding sites with similar stoichiometries (bound dye to monomeric protein molar ratio ∼10), but different association constants (∼6 and 0.1μM(-1)) and ThT quantum yields in fibril-associated state (0.08 and 0.05, respectively). A β-strand-loop-β-strand structural model of 1-83/G26R/W@8 apoA-I fibrils has been proposed, with potential ThT binding sites located in the solvent-exposed grooves of the N-terminal β-sheet layer. Reasoning from the expanded FRET analysis allowing for heterogeneity of ThT binding centers and fibril polymorphism, the most probable locations of high- and low-affinity ThT binding sites were attributed to the grooves T16_Y18 and D20_L22, respectively.

  17. FOXO1 and LXRα downregulate the apolipoprotein A-I gene expression during hydrogen peroxide-induced oxidative stress in HepG2 cells.

    PubMed

    Shavva, Vladimir S; Bogomolova, Alexandra M; Nikitin, Artemy A; Dizhe, Ella B; Oleinikova, Galina N; Lapikov, Ivan A; Tanyanskiy, Dmitry A; Perevozchikov, Andrej P; Orlov, Sergey V

    2017-01-01

    Reactive oxygen species damage various cell components including DNA, proteins, and lipids, and these impairments could be a reason for severe human diseases including atherosclerosis. Forkhead box O1 (FOXO1), an important metabolic transcription factor, upregulates antioxidant and proapoptotic genes during oxidative stress. Apolipoprotein A-I (ApoA-I) forms high density lipoprotein (HDL) particles that are responsible for cholesterol transfer from peripheral tissues to liver for removal in bile in vertebrates. The main sources for plasma ApoA-I in mammals are liver and jejunum. Hepatic apoA-I transcription depends on a multitude of metabolic transcription factors. We demonstrate that ApoA-I synthesis and secretion are decreased during H2O2-induced oxidative stress in human hepatoma cell line HepG2. Here, we first show that FOXO1 binds to site B of apoA-I hepatic enhancer and downregulates apoA-I gene activity in HepG2 cells. Moreover, FOXO1 and LXRα transcription factors participate in H2O2-triggered downregulation of apoA-I gene together with Src, JNK, p38, and AMPK kinase cascades. Mutations of sites B or C as well as the administration of siRNAs against FOXO1 or LXRα to HepG2 cells abolished the hydrogen peroxide-mediated suppression of apoA-I gene.

  18. A fast semi-quantitative LC-MS method for measurement of intact apolipoprotein A-I reveals novel proteoforms in serum.

    PubMed

    Gåfvels, M; Bengtson, P

    2015-03-10

    Surrogate markers for reverse cholesterol transport (RCT) efficiency such as HDL cholesterol and immune methods for apolipoprotein A-I (ApoA-I) may not fully reflect the actual efficiency of the RCT pathway. Several genetic variants and different posttranslational proteoforms of ApoA-I may unevenly affect the functionality of the HDL particle to efflux cholesterol. A method employing top-down immunoaffinity LC-MS of ApoA-I in order to characterize the most prevalent ApoA-I proteoforms in human plasma is described. Diluted plasma was directly injected into a 2D LC-MS system consisting of an affinity column and an analytical column. Enriched ApoA-I fractions were introduced into the MS and intact or fragmented ApoA-I was analyzed. ApoA-I as detected by the described LC-MS method distributes into at least 14 major potential proteoforms exceeding detection limit in human plasma. Substantial amounts of ApoA-I in plasma were found to occur as truncated, oxidized, glycated and glycosylated proteoforms. Levels of glycated ApoA-I distinguished significantly diabetic from non-diabetic samples. In addition novel truncated and glycosylated proteoforms were detected. ApoA-I proteoforms measured by LC-MS represent a useful approach to augment the clinical picture of ApoA-I and its function in health and disease. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. The effect of chromium picolinate on serum cholesterol and apolipoprotein fractions in human subjects.

    PubMed Central

    Press, R. I.; Geller, J.; Evans, G. W.

    1990-01-01

    Chromium has been implicated as a cofactor in the maintenance of normal lipid and carbohydrate metabolism. A deficiency of chromium results from diets low in biologically available chromium. Picolinic acid, a metabolite of tryptophan, forms stable complexes with transitional metal ions, which results in an improved bioavailability of the metal ion chromium. To determine whether or not chromium picolinate is effective in humans, 28 volunteer subjects were given either chromium tripicolinate (3.8 micromol [200 micrograms] chromium) or a placebo daily for 42 days in a double-blind crossover study. A 14-day period off capsules was used between treatments. Levels of total cholesterol, low-density lipoprotein (LDL) cholesterol, and apolipoprotein B, the principal protein of the LDL fraction, decreased significantly while the subjects were ingesting chromium picolinate. The concentration of apolipoprotein A-I, the principal protein of the high-density lipoprotein (HDL) fraction, increased substantially during treatment with chromium picolinate. The HDL-cholesterol level was elevated slightly but not significantly during ingestion of chromium picolinate. Only apolipoprotein B, of the variables measured, was altered significantly during supplementation with the placebo. These observations show that chromium picolinate is efficacious in lowering blood lipids in humans. PMID:2408233

  20. Simvastatin, an HMG-CoA reductase inhibitor, induces the synthesis and secretion of apolipoprotein AI in HepG2 cells and primary hamster hepatocytes.

    PubMed

    Bonn, Victoria; Cheung, Raphael C; Chen, Biao; Taghibiglou, Changiz; Van Iderstine, Stephen C; Adeli, Khosrow

    2002-07-01

    Clinical studies have recently suggested that statin treatment may beneficially elevate plasma concentrations of high density lipoprotein (HDL)-cholesterol in patients with hyperlipidemia. Here, we have investigated the effect of a potent inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase on the synthesis and secretion of apolipoprotein AI (apoAI) in two model systems, HepG2 cells and primary hamster hepatocytes. Cultured cells were incubated with different doses of simvastatin (0.1-10 microM) for a period of 18 h. A dose-dependent increase in synthesis and secretion of apoAI was observed in both cell types. There was a significant increase in the synthesis of apoAI in HepG2 cells (44.3+/-12.1%), and hamster hepatocytes (212+/-2%) after treatment with 10 microM of the statin. The increase in apoAI synthesis appeared to result in a higher level of apoAI secreted into the culture media in both cell types (49.2+/-7.8% in HepG2, 197+/-0.2% in hamster hepatocytes). ApoAI mRNA levels were also significantly increased in both cell types in response to statin treatment. Control experiments with transferrin confirmed specificity of the effect on apoAI secretion. Analysis of a density fraction containing HDL particles in culture media revealed an increase in HDL-associated apoAI of 94.3+/-2.1% in HepG2 cells and 27.0+/-0.03% in hamster hepatocytes following 10 microM simvastatin-treatment. Comparative studies of simvastatin and lovastatin indicated a differential ability to induce apoAI synthesis and secretion, with simvastatin having a more significant effect. Thus, acute statin treatment of cultured hepatocytes (transformed as well as primary) resulted in a significant upregulation of apoAI mRNA and apoAI synthesis, causing oversecretion of apoAI and HDL extracellularly. The stimulatory effect on apoAI synthesis and secretion may thus explain the clinical observation of an elevated plasma HDL-cholesterol level in hyperlipidemic patients treated with

  1. Decreased apolipoprotein A-I level indicates poor prognosis in extranodal natural killer/T-cell lymphoma, nasal type

    PubMed Central

    Quan, Qi; Chen, Qi; Chen, Ping; Jiang, Li; Li, Tingwei; Qiu, Huijuan; Zhang, Bei

    2016-01-01

    Background Extranodal natural killer (NK)/T-cell lymphoma, nasal type (ENKTL) is an invasive lymphoid malignancy with unfavorable survival, for which a prognostic model has not yet been validated. We hypothesized that serum apolipoprotein A-I (ApoA-I) may serve as a novel prognostic marker for ENKTL. Patients and methods A total of 236 newly diagnosed cases of ENKTL were analyzed retrospectively. Results The optimal cutoff value for the serum ApoA-I level was determined to be 0.95 g/L. A total of 154 and 82 cases were assigned to the high and low ApoA-I groups, respectively. Patients in the low ApoA-I group tended to present with poorer clinical features, a lower complete remission rate (P=0.001), and poor median progression-free survival (P<0.001) and overall survival (P<0.001). Multivariate analysis using Cox model showed that the serum ApoA-I level was an independent prognostic marker of overall survival (P<0.001) and progression-free survival (P<0.001) for ENKTL patients. For cases in the low-risk group, as assessed by International Prognostic Index, Prognosis Index for peripheral T-cell lymphoma, unspecified, and Korean Prognostic Index, the serum ApoA-I level was able to differentiate cases with poor outcomes from cases with good outcomes. Conclusion Our results showed that the baseline serum ApoA-I level was helpful for predicting ENKTL prognosis. PMID:27051293

  2. Deficiency in apolipoprotein A-I ablates the pharmacological effects of metformin on plasma glucose homeostasis and hepatic lipid deposition.

    PubMed

    Karavia, Eleni A; Hatziri, Aikaterini; Kalogeropoulou, Christina; Papachristou, Nikolaos I; Xepapadaki, Eva; Constantinou, Caterina; Natsos, Anastasios; Petropoulou, Peristera-Ioanna; Sasson, Shlomo; Papachristou, Dionysios J; Kypreos, Kyriakos E

    2015-11-05

    Recently, we showed that deficiency in apolipoprotein A-I (ApoA-I) sensitizes mice to diet-induced obesity, glucose intolerance and NAFLD. Here we investigated the potential involvement of ApoA-I in the pharmacological effects of metformin on glucose intolerance and NAFLD development. Groups of apoa1-deficient (apoa1(-/-)) and C57BL/6 mice fed western-type diet were either treated with a daily dose of 300 mg/kg metformin for 18 weeks or left untreated for the same period. Then, histological and biochemical analyses were performed. Metformin treatment led to a comparable reduction in plasma insulin levels in both C57BL/6 and apoa1(-/-) mice following intraperitoneal glucose tolerance test. However, only metformin-treated C57BL/6 mice maintained sufficient peripheral insulin sensitivity to effectively clear glucose following the challenge, as indicated by a [(3)H]-2-deoxy-D-glucose uptake assay in isolated soleus muscle. Similarly, deficiency in ApoA-I ablated the effect of metformin on hepatic lipid deposition and NAFLD development. Gene expression analysis indicated that the effects of ApoA-I on metformin treatment may be independent of adenosine monophosphate-activated protein kinase (AMPK) activation and de novo lipogenesis. Interestingly, metformin treatment reduced mitochondrial oxidative phosphorylation function only in apoa1(-/-) mice. Our data show that the role of ApoA-I in diabetes extends to the modulation of the pharmacological actions of metformin, a common drug for the treatment of type 2 diabetes. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Inflammatory markers modify the risk of recurrent coronary events associated with apolipoprotein A-I in postinfarction patients.

    PubMed

    Wang, Meng; Corsetti, James; McNitt, Scott; Rich, David Q; Sparks, Charles E; Moss, Arthur J; Zareba, Wojciech

    Laboratory findings have suggested that systemic and vascular inflammation can impair the antiatherogenic function of high-density lipoproteins (HDLs). However, evidence from population studies is sparse. The objective of the study was to assess if blood inflammatory markers modify the risk of recurrent coronary events associated with apolipoprotein A-I (apoA-I) and HDL cholesterol (HDL-C) among postinfarction patients. ApoA-I, HDL-C, and inflammatory markers (C-reactive protein [CRP], serum amyloid A (SAA), fibrinogen, von Willebrand factor [vWF], and D-dimer) were measured from blood samples of 1028 patients drawn 2 months after an index myocardial infarction (MI). Patients were followed up for the composite coronary endpoint (nonfatal MI, coronary death, or unstable angina) for an average of 26 months. Cox proportional hazard models were used to assess effect modifications for the association of apoA-I and HDL-C with coronary risk by each inflammatory marker. CRP significantly modified the risk of recurrent coronary events associated with apoA-I. Among the entire population, multivariable-adjusted hazard ratios associated with each standard deviation increase in apoA-I for those with low and high CRP levels were 0.89 and 1.35, respectively (P value for interaction = .008). vWF was a significant effect modifier of the apoA-I/coronary risk association only among diabetic patients (hazard ratios were 0.56 and 1.43, for diabetic patients with low and high vWF levels, respectively; P value for interaction = .002). No effect modification was observed for the HDL-C/coronary risk association. Among stable post-MI patients, CRP modified the risk of recurrent coronary events associated with apoA-I. VWF modified this association only among the diabetic subgroup. Copyright © 2017 National Lipid Association. Published by Elsevier Inc. All rights reserved.

  4. Mycoplasma gallisepticum (HS strain) surface lipoprotein pMGA interacts with host apolipoprotein A-I during infection in chicken.

    PubMed

    Hu, Fuli; Zhao, Chengcheng; Bi, Dingren; Tian, Wei; Chen, Jiao; Sun, Jianjun; Peng, Xiuli

    2016-02-01

    The adhesin protein from Mycoplasma gallisepticum (HS strain), namely pMGA1.2, is required for M. gallisepticum (MG) infection in chicken. However, the host factor(s) that interact with pMGA1.2 is not known. In this study, we prepared the membrane fraction of trachea epithelial cells from chicken embryos. Using an improved virus overlay protein blot assay (VOPBA) and glutathione S-transferase (GST) pull-down assay, we found that pMGA1.2 specifically bound to a ∼30 kDa host protein. This host protein was further identified by mass spectrometry as chicken apolipoprotein A-I (ApoA-I). We expressed and purified the recombinant ApoA-I protein in Escherichia coli and confirmed that it bound to the purified pMGA1.2 protein in vitro. Transiently expressed pMGA1.2 and ApoA-I were colocalized in HeLa cells. Finally, we designed small interfering RNA (siRNA) molecules to knock down the expression of either ApoA-I or pMGA1.2, which inhibited the MG-induced cell cycle disruption in cells of chicken embryo fibroblast cell line (DF-1). Similarly, knockdown of ApoA-I inhibited the cilia loss and damage in chicken trachea cells in MG infection. In summary, ApoA-I may be an essential host factor in MG infection through interacting with pMGA1.2.

  5. The A's Have It: Developing Apolipoprotein A-I Mimetic Peptides Into a Novel Treatment for Asthma.

    PubMed

    Yao, Xianglan; Gordon, Elizabeth M; Barochia, Amisha V; Remaley, Alan T; Levine, Stewart J

    2016-08-01

    New treatments are needed for patients with asthma who are refractory to standard therapies, such as individuals with a phenotype of "type 2-low" inflammation. This important clinical problem could potentially be addressed by the development of apolipoprotein A-I (apoA-I) mimetic peptides. ApoA-I interacts with its cellular receptor, the ATP-binding cassette subfamily A, member 1 (ABCA1), to facilitate cholesterol efflux out of cells to form nascent high-density lipoprotein particles. The ability of the apoA-I/ABCA1 pathway to promote cholesterol efflux from cells that mediate adaptive immunity, such as antigen-presenting cells, can attenuate their function. Data from experimental murine models have shown that the apoA-I/ABCA1 pathway can reduce neutrophilic airway inflammation, primarily by suppressing the production of granulocyte-colony stimulating factor. Furthermore, administration of apoA-I mimetic peptides to experimental murine models of allergic asthma has decreased both neutrophilic and eosinophilic airway inflammation, as well as airway hyperresponsiveness and mucous cell metaplasia. Higher serum levels of apoA-I have also been associated with less severe airflow obstruction in patients with asthma. Collectively, these results suggest that the apoA-I/ABCA1 pathway may have a protective effect in asthma, and support the concept of advancing inhaled apoA-I mimetic peptides to clinical trials that can assess their safety and effectiveness. Thus, we propose that the development of inhaled apoA-I mimetic peptides as a new treatment could represent a clinical advance for patients with severe asthma who are unresponsive to other therapies. Published by Elsevier Inc.

  6. Genome-wide screen for modulation of hepatic apolipoprotein A-I (ApoA-I) secretion.

    PubMed

    Miles, Rebecca R; Perry, William; Haas, Joseph V; Mosior, Marian K; N'Cho, Mathias; Wang, Jian W J; Yu, Peng; Calley, John; Yue, Yong; Carter, Quincy; Han, Bomie; Foxworthy, Patricia; Kowala, Mark C; Ryan, Timothy P; Solenberg, Patricia J; Michael, Laura F

    2013-03-01

    Control of plasma cholesterol levels is a major therapeutic strategy for management of coronary artery disease (CAD). Although reducing LDL cholesterol (LDL-c) levels decreases morbidity and mortality, this therapeutic intervention only translates into a 25-40% reduction in cardiovascular events. Epidemiological studies have shown that a high LDL-c level is not the only risk factor for CAD; low HDL cholesterol (HDL-c) is an independent risk factor for CAD. Apolipoprotein A-I (ApoA-I) is the major protein component of HDL-c that mediates reverse cholesterol transport from tissues to the liver for excretion. Therefore, increasing ApoA-I levels is an attractive strategy for HDL-c elevation. Using genome-wide siRNA screening, targets that regulate hepatocyte ApoA-I secretion were identified through transfection of 21,789 siRNAs into hepatocytes whereby cell supernatants were assayed for ApoA-I. Approximately 800 genes were identified and triaged using a convergence of information, including genetic associations with HDL-c levels, tissue-specific gene expression, druggability assessments, and pathway analysis. Fifty-nine genes were selected for reconfirmation; 40 genes were confirmed. Here we describe the siRNA screening strategy, assay implementation and validation, data triaging, and example genes of interest. The genes of interest include known and novel genes encoding secreted enzymes, proteases, G-protein-coupled receptors, metabolic enzymes, ion transporters, and proteins of unknown function. Repression of farnesyltransferase (FNTA) by siRNA and the enzyme inhibitor manumycin A caused elevation of ApoA-I secretion from hepatocytes and from transgenic mice expressing hApoA-I and cholesterol ester transfer protein transgenes. In total, this work underscores the power of functional genetic assessment to identify new therapeutic targets.

  7. Cholesterol Independent Suppression of Lymphocyte Activation, Autoimmunity and Glomerulonephritis by Apolipoprotein A-I in Normocholesterolemic Lupus-Prone Mice

    PubMed Central

    Black, Leland L.; Srivastava, Roshni; Schoeb, Trenton R.; Moore, Ray D.; Barnes, Stephen; Kabarowski, Janusz H.

    2015-01-01

    Apolipoprotein A-I (ApoA-I), the major lipid-binding protein of high-density lipoprotein (HDL), can prevent autoimmunity and suppress inflammation in hypercholesterolemic mice by attenuating lymphocyte cholesterol accumulation and removing tissue oxidized lipids. However, whether ApoA-I mediates immune suppressive or anti-inflammatory effects in normocholesterolemic conditions and the mechanisms involved remain unresolved. We transferred bone marrow from SLE-prone Sle123 mice into normal, ApoA-I knockout (ApoA-I−/−) and ApoA-I transgenic (ApoA-Itg) mice. Increased ApoA-I in ApoA-Itg mice suppressed CD4+ T and B cell activation without changing lymphocyte cholesterol levels or reducing major ApoA-I-binding oxidized fatty acids. Unexpectedly, oxidized fatty acid peroxisome proliferator-activated receptor gamma (PPARγ) ligands 13-hydroxyoctadecadienoic acid (HODE) and 9-HODE were increased in lymphocytes of autoimmune ApoA-Itg mice. ApoA-I reduced Th1 cells independently of changes in CD4+FoxP3+ regulatory T cells or CD11c+ dendritic cell activation and migration. Follicular helper T cells, germinal center B cells and autoantibodies were also lower in ApoA-Itg mice. Transgenic ApoA-I also improved SLE-mediated glomerulonephritis. However, ApoA-I deficiency did not have opposite effects on autoimmunity or glomerulonephritis, possibly due to compensatory increases of ApoE on HDL. We conclude that although compensatory mechanisms prevent pro-inflammatory effects of ApoA-I deficiency in normocholesterolemic mice, increasing ApoA-I can attenuate lymphocyte activation and autoimmunity in SLE independently of cholesterol transport, possibly through oxidized fatty acid PPARγ ligands, and can reduce renal inflammation in glomerulonephritis. PMID:26466956

  8. Enthalpy-driven apolipoprotein A-I and lipid bilayer interaction indicating protein penetration upon lipid binding.

    PubMed

    Arnulphi, Cristina; Jin, Lihua; Tricerri, M Alejandra; Jonas, Ana

    2004-09-28

    The interaction of lipid-free apolipoprotein A-I (apoA-I) with small unilamellar vesicles (SUVs) of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) with and without free cholesterol (FC) was studied by isothermal titration calorimetry and circular dichroism spectroscopy. Parameters reported are the affinity constant (K(a)), the number of protein molecules bound per vesicle (n), enthalpy change (DeltaH degrees), entropy change (DeltaS degrees ), and the heat capacity change (DeltaC(p) degrees). The binding process of apoA-I to SUVs of POPC plus 0-20% (mole) FC was exothermic between 15 and 37 degrees C studied, accompanied by a small negative entropy change, making enthalpy the main driving force of the interaction. The presence of cholesterol in the vesicles increased the binding affinity and the alpha-helix content of apoA-I but lowered the number of apoA-I bound per vesicle and the enthalpy and entropy changes per bound apoA-I. Binding affinity and stoichiometry were essentially invariant of temperature for binding to SUVs of POPC/FC at a molar ratio of 6/1 at (2.8-4) x 10(6) M(-1) and 2.4 apoA-I molecules bound per vesicle or 1.4 x 10(2) phospholipids per bound apoA-I. A plot of DeltaH degrees against temperature displayed a linear behavior, from which the DeltaC(p) degrees per mole of bound apoA-I was calculated to be -2.73 kcal/(mol x K). These results suggested that binding of apoA-I to POPC vesicles is characterized by nonclassical hydrophobic interactions, with alpha-helix formation as the main driving force for the binding to cholesterol-containing vesicles. In addition, comparison to literature data on peptides suggested a cooperativity of the helices in apoA-I in lipid interaction.

  9. Conservation of apolipoprotein A-I's central domain structural elements upon lipid association on different high-density lipoprotein subclasses.

    PubMed

    Oda, Michael N; Budamagunta, Madhu S; Geier, Ethan G; Chandradas, Sajiv H; Shao, Baohai; Heinecke, Jay W; Voss, John C; Cavigiolio, Giorgio

    2013-10-01

    The antiatherogenic properties of apolipoprotein A-I (apoA-I) are derived, in part, from lipidation-state-dependent structural elements that manifest at different stages of apoA-I's progression from lipid-free protein to spherical high-density lipoprotein (HDL). Previously, we reported the structure of apoA-I's N-terminus on reconstituted HDLs (rHDLs) of different sizes. We have now investigated at the single-residue level the conformational adaptations of three regions in the central domain of apoA-I (residues 119-124, 139-144, and 164-170) upon apoA-I lipid binding and HDL formation. An important function associated with these residues of apoA-I is the activation of lecithin:cholesterol acyltransferase (LCAT), the enzyme responsible for catalyzing HDL maturation. Structural examination was performed by site-directed tryptophan fluorescence and spin-label electron paramagnetic resonance spectroscopies for both the lipid-free protein and rHDL particles 7.8, 8.4, and 9.6 nm in diameter. The two methods provide complementary information about residue side chain mobility and molecular accessibility, as well as the polarity of the local environment at the targeted positions. The modulation of these biophysical parameters yielded new insight into the importance of structural elements in the central domain of apoA-I. In particular, we determined that the loosely lipid-associated structure of residues 134-145 is conserved in all rHDL particles. Truncation of this region completely abolished LCAT activation but did not significantly affect rHDL size, reaffirming the important role of this structural element in HDL function.

  10. Extended-release niacin alters the metabolism of plasma apolipoprotein (apo) A-I- and apoB-containing lipoproteins

    PubMed Central

    Lamon-Fava, Stefania; Diffenderfer, Margaret R.; Barrett, P. Hugh R.; Buchsbaum, Aaron; Nyaku, Mawuli; Horvath, Katalin V.; Asztalos, Bela F.; Otokozawa, Seiko; Ai, Masumi; Matthan, Nirupa R.; Lichtenstein, Alice H; Dolnikowski, Gregory G.; Schaefer, Ernst J.

    2009-01-01

    Objectives Extended-release niacin effectively lowers plasma TG levels and raises plasma HDL cholesterol levels, but the mechanisms responsible for these effects are unclear. Methods and Results We examined the effects of extended-release niacin (2 g/d) and extended-release niacin (2 g/d) plus lovastatin (40 mg/d), relative to placebo, on the kinetics of apolipoprotein (apo) A-I and apoA-II in HDL, apoB-100 in TG-rich lipoproteins (TRL), intermediate-density lipoproteins (IDL) and LDL, and apoB-48 in TRL in five men with combined hyperlipidemia. Niacin significantly increased HDL cholesterol and apoA-I concentrations, associated with a significant increase in apoA-I production rate (PR) and no change in fractional catabolic rate (FCR). Plasma TRL apoB-100 levels were significantly lowered by niacin, accompanied by a trend toward an increase in FCR and no change in PR. Niacin treatment significantly increased TRL apoB-48 FCR but had no effect on apoB-48 PR. No effects of niacin on concentrations or kinetic parameters of IDL and LDL apoB-100 and HDL apoA-II were noted. The addition of lovastatin to niacin promoted a lowering in LDL apoB-100 due to increased LDL apoB-100 FCR. Conclusion Niacin treatment was associated with significant increases in HDL apoA-I concentrations and production, as well as enhanced clearance of TRL apoB-100 and apoB-48. PMID:18566298

  11. Apolipoprotein A-I, A-II, and H mRNA and protein accumulation sites in the developing lung in late gestation

    PubMed Central

    2011-01-01

    Background Expression of apolipoprotein A-I (apoA-I), A-II, and H was previously observed at 16 to 50-fold higher levels in the fetal than the adult mouse lung. Here, sites of apoA-I, A-II, and H mRNA and protein accumulation were determined in mouse fetal lungs by in situ hybridization and immunohistochemistry in late gestation. Results Expression sites vary for the three genes and change for the distal epithelium before the end of the canalicular stage, thus where and when the surge of surfactant synthesis occurs. Messenger of apoH, but not those of apoA-I and A-II, was also observed in the proximal epithelium and smooth muscles surrounding arteries. In contrast to apoC-II protein, none of the three studied apolipoproteins accumulated within secretory granule-like structures. Immunohistochemistry revealed that apoA-I and apoH accumulated mainly in capillaries. Three different positive signals with the anti-apoA-II antibody were found: one transient signal in the nucleus of a portion of mesenchymal cells, a second at lower levels throughout the mesenchyme, and another in capillaries with a specific increase from gestation day 17.5/18.5. Conclusion Temporal and geographic co-expression of apoAI, AII, and H genes with surfactant production site suggests that the three apolipoproteins are secreted to play roles supporting the lung-specific surfactant lipid-related metabolism. PMID:21756353

  12. Apolipoprotein A-I mimetic peptide 4F blocks sphingomyelinase-induced LDL aggregation.

    PubMed

    Nguyen, Su Duy; Javanainen, Matti; Rissanen, Sami; Zhao, Hongxia; Huusko, Jenni; Kivelä, Annukka M; Ylä-Herttuala, Seppo; Navab, Mohamad; Fogelman, Alan M; Vattulainen, Ilpo; Kovanen, Petri T; Öörni, Katariina

    2015-06-01

    Lipolytic modification of LDL particles by SMase generates LDL aggregates with a strong affinity for human arterial proteoglycans and may so enhance LDL retention in the arterial wall. Here, we evaluated the effects of apoA-I mimetic peptide 4F on structural and functional properties of the SMase-modified LDL particles. LDL particles with and without 4F were incubated with SMase, after which their aggregation, structure, and proteoglycan binding were analyzed. At a molar ratio of L-4F to apoB-100 of 2.5 to 20:1, 4F dose-dependently inhibited SMase-induced LDL aggregation. At a molar ratio of 20:1, SMase-induced aggregation was fully blocked. Binding of 4F to LDL particles inhibited SMase-induced hydrolysis of LDL by 10% and prevented SMase-induced LDL aggregation. In addition, the binding of the SMase-modified LDL particles to human aortic proteoglycans was dose-dependently inhibited by pretreating LDL with 4F. The 4F stabilized apoB-100 conformation and inhibited SMase-induced conformational changes of apoB-100. Molecular dynamic simulations showed that upon binding to protein-free LDL surface, 4F locally alters membrane order and fluidity and induces structural changes to the lipid layer. Collectively, 4F stabilizes LDL particles by preventing the SMase-induced conformational changes in apoB-100 and so blocks SMase-induced LDL aggregation and the resulting increase in LDL retention. Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.

  13. Effects of dietary fat amount and saturation on the regulation of hepatic mRNA and plasma apolipoprotein A-I in rats.

    PubMed

    Calleja, L; Trallero, M C; Carrizosa, C; Méndez, M T; Palacios-Alaiz, E; Osada, J

    2000-09-01

    The effects of the amount of dietary fat and saturation together with cholesterol both on hepatic apolipoprotein A-I gene mRNA levels and on plasma levels of this apolipoprotein were studied in male rats. To achieve these goals, seven groups of male Wistar rats were established: control group (n=5) consuming chow diet; cholesterol group (n=4) fed on a chow diet containing 0.1% (w/w) cholesterol; coco group (n=5) fed on a chow diet containing 0.1% (w/w) cholesterol and 40% coconut oil; corn group (n=5) fed on a chow diet containing 0.1% (w/w) cholesterol and 40% corn oil; and three olive groups consuming a chow diet containing 0.1% (w/w) cholesterol and percentages of 5 (n=5), 10 (n=4) and 40% (n=5), respectively, of olive oil. Animals were kept on these diets for 2 months and then sacrificed for lipoprotein, apolipoprotein and hepatic mRNA analysis. Dietary cholesterol by itself was hypercholesterolemic when compared to chow diet, an effect that was mainly due to an increase in LDL-cholesterol. Corn oil had a hypocholesterolemic action, whether compared to chow or to cholesterol diet, due to a reduction in HDL-cholesterol as well as LDL-cholesterol. HDL-cholesterol levels of 40% olive oil diet were lower than those corresponding to coconut oil and higher than those found in corn oil diet. When compared to control or cholesterol diets, plasma apoA-I concentration appeared significantly increased in coconut and 40% olive oil diets. Coconut oil or corn oil diets did not induce any significant change in apoA-I mRNA compared to control or cholesterol diets. Compared to cholesterol diet, 40 and 10% olive oil diets induced a significant increase in the expression of this message. A positive and significant (r=0.97, P<0.01) correlation between plasma apolipoprotein A-I concentration and its hepatic mRNA, was observed when the amount of dietary olive oil was 40% (w/w). A significant negative (r=-0.97, P<0.01) correlation was found in the corn oil group and no significant

  14. PPARγ Represses Apolipoprotein A-I Gene but Impedes TNFα-Mediated ApoA-I Downregulation in HepG2 Cells.

    PubMed

    Shavva, Vladimir S; Mogilenko, Denis A; Bogomolova, Alexandra M; Nikitin, Artemy A; Dizhe, Ella B; Efremov, Alexander M; Oleinikova, Galina N; Perevozchikov, Andrej P; Orlov, Sergey V

    2016-09-01

    Apolipoprotein A-I (ApoA-I) is the main anti-atherogenic component of human high-density lipoproteins (HDL). ApoA-I gene expression is regulated by several nuclear receptors, which are the sensors for metabolic changes during development of cardiovascular diseases. Activation of nuclear receptor PPARγ has been shown to impact lipid metabolism as well as inflammation. Here, we have shown that synthetic PPARγ agonist GW1929 decreases both ApoA-I mRNA and protein levels in HepG2 cells and the effect of GW1929 on apoA-I gene transcription depends on PPARγ. PPARγ binds to the sites A and C within the hepatic enhancer of apoA-I gene and the negative regulation of apoA-I gene transcription by PPARγ appears to be realized via the site C (-134 to -119). Ligand activation of PPARγ leads to an increase of LXRβ and a decrease of PPARα binding to the apoA-I gene hepatic enhancer in HepG2 cells. GW1929 abolishes the TNFα-mediated decrease of ApoA-I mRNA expression in both HepG2 and Caco-2 cells but does not block TNFα-mediated inhibition of ApoA-I protein secretion by HepG2 cells. These data demonstrate that complex of PPARγ with GW1929 is a negative regulator involved in the control of ApoA-I expression and secretion in human hepatocyte- and enterocyte-like cells. J. Cell. Biochem. 117: 2010-2022, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  15. Apolipoprotein A-I Modulates Processes Associated with Diet-Induced Nonalcoholic Fatty Liver Disease in Mice

    PubMed Central

    Karavia, Eleni A; Papachristou, Dionysios J; Liopeta, Kassiani; Triantaphyllidou, Irene-Eva; Dimitrakopoulos, Odyssefs; Kypreos, Kyriakos E

    2012-01-01

    Apolipoprotein A-I (apoA-I) is the main protein of high-density lipoprotein (HDL). We investigated the involvement of apoA-I in diet-induced accumulation of triglycerides in hepatocytes and its potential role in the treatment of nonalcoholic fatty liver disease (NAFLD). ApoA-I–deficient (apoA-I−/−) mice showed increased diet-induced hepatic triglyceride deposition and disturbed hepatic histology while they exhibited reduced glucose tolerance and insulin sensitivity. Quantification of FASN (fatty acid synthase 1), DGAT-1 (diacylglycerol O-acyltransferase 1), and PPARγ (peroxisome proliferator-activated receptor γ) mRNA expression suggested that the increased hepatic triglyceride content of the apoA-I−/− mice was not due to de novo synthesis of triglycerides. Similarly, metabolic profiling did not reveal differences in the energy expenditure between the two mouse groups. However, apoA-I−/− mice exhibited enhanced intestinal absorption of dietary triglycerides (3.6 ± 0.5 mg/dL/min for apoA-I−/− versus 2.0 ± 0.7 mg/dL/min for C57BL/6 mice, P < 0.05), accelerated clearance of postprandial triglycerides and a reduced rate of hepatic very low density lipoprotein (VLDL) triglyceride secretion (9.8 ± 1.1 mg/dL/min for apoA-I−/− versus 12.5 ± 1.3 mg/dL/min for C57BL/6 mice, P < 0.05). In agreement with these findings, adenovirus-mediated gene transfer of apoA-IMilano in apoA-I−/− mice fed a Western-type diet for 12 wks resulted in a significant reduction in hepatic triglyceride content and an improvement of hepatic histology and architecture. Our data extend the current knowledge on the functions of apoA-I, indicating that in addition to its well-established properties in atheroprotection, it is also an important modulator of processes associated with diet-induced hepatic lipid deposition and NAFLD development in mice. Our findings raise the interesting possibility that expression of therapeutic forms of apoA-I by gene therapy approaches may

  16. Apolipoprotein A-I exchange is impaired in metabolic syndrome patients asymptomatic for diabetes and cardiovascular disease.

    PubMed

    Borja, Mark S; Hammerson, Bradley; Tang, Chongren; Savinova, Olga V; Shearer, Gregory C; Oda, Michael N

    2017-01-01

    We tested the hypothesis that HDL-apolipoprotein A-I exchange (HAE), a measure of high-density lipoprotein (HDL) function and a key step in reverse cholesterol transport (RCT), is impaired in metabolic syndrome (MetSyn) patients who are asymptomatic for diabetes and cardiovascular disease. We also compared HAE with cell-based cholesterol efflux capacity (CEC) to address previous reports that CEC is enhanced in MetSyn populations. HAE and ABCA1-specific CEC were measured as tests of HDL function in 60 MetSyn patients and 14 normolipidemic control subjects. Predictors of HAE and CEC were evaluated with multiple linear regression modeling using clinical markers of MetSyn and CVD risk. HAE was significantly reduced in MetSyn patients (49.0 ± 10.9% vs. 61.2 ± 6.1%, P < 0.0001), as was ABCA1-specific CEC (10.1 ± 1.6% vs. 12.3 ± 2.0%, P < 0.002). Multiple linear regression analysis identified apoA-I concentration as a significant positive predictor of HAE, and MetSyn patients had significantly lower HAE per mg/dL of apoA-I (P = 0.004). MetSyn status was a negative predictor of CEC, but triglyceride (TG) was a positive predictor of CEC, with MetSyn patients having higher CEC per mg/dL of TG, but lower overall CEC compared to controls. MetSyn patients have impaired HAE that contributes to reduced capacity for ABCA1-mediated CEC. MetSyn status is inversely correlated with CEC but positively correlated with TG, which explains the contradictory results from earlier MetSyn studies focused on CEC. HAE and CEC are inhibited in MetSyn patients over a broad range of absolute apoA-I and HDL particle levels, supporting the observation that this patient population bears significant residual cardiovascular disease risk.

  17. Apolipoprotein A-I exchange is impaired in metabolic syndrome patients asymptomatic for diabetes and cardiovascular disease

    PubMed Central

    Borja, Mark S.; Hammerson, Bradley; Tang, Chongren; Savinova, Olga V.; Shearer, Gregory C.

    2017-01-01

    Objective We tested the hypothesis that HDL-apolipoprotein A-I exchange (HAE), a measure of high-density lipoprotein (HDL) function and a key step in reverse cholesterol transport (RCT), is impaired in metabolic syndrome (MetSyn) patients who are asymptomatic for diabetes and cardiovascular disease. We also compared HAE with cell-based cholesterol efflux capacity (CEC) to address previous reports that CEC is enhanced in MetSyn populations. Methods HAE and ABCA1-specific CEC were measured as tests of HDL function in 60 MetSyn patients and 14 normolipidemic control subjects. Predictors of HAE and CEC were evaluated with multiple linear regression modeling using clinical markers of MetSyn and CVD risk. Results HAE was significantly reduced in MetSyn patients (49.0 ± 10.9% vs. 61.2 ± 6.1%, P < 0.0001), as was ABCA1-specific CEC (10.1 ± 1.6% vs. 12.3 ± 2.0%, P < 0.002). Multiple linear regression analysis identified apoA-I concentration as a significant positive predictor of HAE, and MetSyn patients had significantly lower HAE per mg/dL of apoA-I (P = 0.004). MetSyn status was a negative predictor of CEC, but triglyceride (TG) was a positive predictor of CEC, with MetSyn patients having higher CEC per mg/dL of TG, but lower overall CEC compared to controls. Conclusions MetSyn patients have impaired HAE that contributes to reduced capacity for ABCA1-mediated CEC. MetSyn status is inversely correlated with CEC but positively correlated with TG, which explains the contradictory results from earlier MetSyn studies focused on CEC. HAE and CEC are inhibited in MetSyn patients over a broad range of absolute apoA-I and HDL particle levels, supporting the observation that this patient population bears significant residual cardiovascular disease risk. PMID:28767713

  18. High-density lipoprotein 3 and apolipoprotein A-I alleviate platelet storage lesion and release of platelet extracellular vesicles.

    PubMed

    Pienimaeki-Roemer, Annika; Fischer, Astrid; Tafelmeier, Maria; Orsó, Evelyn; Konovalova, Tatiana; Böttcher, Alfred; Liebisch, Gerhard; Reidel, Armin; Schmitz, Gerd

    2014-09-01

    Stored platelet (PLT) concentrates (PLCs) for transfusion develop a PLT storage lesion (PSL), decreasing PLT viability and function with profound lipidomic changes and PLT extracellular vesicle (PL-EV) release. High-density lipoprotein 3 (HDL3 ) improves PLT homeostasis through silencing effects on PLT activation in vivo. This prompted us to investigate HDL3 and apolipoprotein A-I (apoA-I) as PSL-antagonizing agents. Healthy donor PLCs were split into low-volume standard PLC storage bags and incubated with native (n)HDL3 or apoA-I from plasma ethanol fractionation (precipitate IV) for 5 days under standard blood banking conditions. Flow cytometry, Born aggregometry, and lipid mass spectrometry were carried out to analyze PL-EV release, PLT aggregation, agonist-induced PLT surface marker expression, and PLT and plasma lipid compositions. Compared to control, added nHDL3 and apoA-I significantly reduced PL-EV release by up to -62% during 5 days, correlating with the added apoA-I concentration. At the lipid level, nHDL3 and apoA-I antagonized PLT lipid loss (+12%) and decreased cholesteryl ester (CE)/free cholesterol (FC) ratios (-69%), whereas in plasma polyunsaturated/saturated CE ratios increased (+3%) and CE 16:0/20:4 ratios decreased (-5%). Administration of nHDL3 increased PLT bis(monoacylglycero)phosphate/phosphatidylglycerol (+102%) and phosphatidic acid/lysophosphatidic acid (+255%) ratios and improved thrombin receptor-activating peptide 6-induced PLT aggregation (+5%). nHDL3 and apoA-I improve PLT membrane homeostasis and intracellular lipid processing and increase CE efflux, antagonizing PSL-related reduction in PLT viability and function and PL-EV release. We suggest uptake and catabolism of nHDL3 into the PLT open canalicular system. As supplement in PLCs, nHDL3 or apoA-I from Fraction IV of plasma ethanol fractionation have the potential to improve PLC quality to prolong storage. © 2014 AABB.

  19. Effect of apolipoprotein a-I complex with tetrahydrocortisone on protein biosynthesis and glucose absorption by rat hepatocytes.

    PubMed

    Sumenkova, D V; Knyazev, R A; Guschya, R S; Polyakov, L M; Panin, L E

    2009-08-01

    We studied the effect of apolipoprotein A-I-tetrahydrocortisone complex on (14)C glucose absorption and lactate accumulation and on the rate of protein biosynthesis in isolated rat hepatocytes. The presence of apolipoprotein A-I-tetrahydrocortisone complex in the incubation medium increased absorption of labeled glucose by hepatocytes by 52%, while lactate content in the conditioning medium increased 4-fold. The rate of protein biosynthesis increased by 80% in comparison with control cells. It is hypothesized that the increase in protein biosynthesis rate in hepatocytes under the effect of apolipoprotein A-I-tetrahydrocortisone complex is due to stimulation of energy metabolism, specifically, of its glycolytic component.

  20. Association of a DNA polymorphism of the apolipoprotein AI-CIII-AIV gene cluster with myocardial infarction in a Tunisian population.

    PubMed

    Sediri, Yousra; Kallel, Amani; Feki, Moncef; Mourali, Sami; Elasmi, Monia; Abdessalem, Salem; Mechmeche, Rachid; Jemaa, Riadh; Kaabachi, Naziha

    2011-08-01

    Apolipoproteins AI-CIII-AIV play important roles in the metabolism of triglycerides and high-density lipoprotein cholesterol. However, whether genetic variations in the ApoAI-CIII-AIV gene cluster are associated with the risk of myocardial infarction (MI) remains uncertain. In the present study, we examined a possible association of the ApoCIII SacI polymorphism in the ApoAI-CIII-AIV gene cluster with lipid parameters and MI in a sample of the Tunisian population. A total of 326 Tunisian patients with MI and 361 controls were included in the study. Genotypes were determined by polymerase chain reaction--restriction fragment length polymorphism (PCR-RFLP) analysis. A significant difference in genotype distribution and allele frequency was observed between patients and controls. At the multivariate analysis after adjustment for traditional vascular risk factors, the ApoCIII SacI polymorphism was significantly associated with MI, according to co-dominant and dominant models (co-dominant model odds ratio [OR]: 1.53, 95% confidence interval [CI]: 1.0-2.35, p=0.04; dominant model OR: 2.02, 95% CI: 1.11-3.67, p=0.02). The MI patient group showed a significant higher frequency of the S2 allele compared to the controls (10.2% vs. 6.5%; OR: 1.64, 95% CI: 1.10-2.47, p=0.01). There was no statistically significant association between ApoAI-CIII-AIV cluster gene polymorphism and lipid, lipoprotein, and apolipoprotein levels in both MI patients and controls. In the current study, a significant association between the ApoCIII SacI polymorphism (presence of S2 allele) and MI in the Tunisian population was found. Copyright © 2011 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.

  1. A novel compound 4010B-30 upregulates apolipoprotein A-I gene expression through activation of PPARγ in HepG2 cells.

    PubMed

    Du, Yu; Wang, Li; Si, Shuyi; Yang, Yuan; Hong, Bin

    2015-04-01

    Apolipoprotein (Apo) A-I is the major lipoprotein content of HDL and upregulating endogenous ApoA-I expression has been proposed as a desirable approach to raise the functional HDL. In this study we investigated the effect of a novel small molecule 4010B-30 on transcriptional regulation of ApoA-I gene in HepG2 cells, and the influence on the level of ApoA-I expression and function. Then the mechanisms by which 4010B-30 regulated ApoA-I expression was further explored. In human hepatic HepG2 cells, 4010B-30 increased the mRNA level and the protein production of ApoA-I both in cell lysates and media. The 4010B-30-induced ApoA-I containing particles increased cholesterol efflux from RAW264.7 macrophages. 4010B-30 also upregulated ABCA1 expression confirmed by transcriptional activity assay and Western blot analysis in both HepG2 and RAW264.7 cells. Promoter luciferase assay was used to identify the 4010B-30-responsive region which is mapped to the proximal -277bp region of the ApoA-I promoter. Further study indicated that the regulation of 4010B-30 on ApoA-I transcription or protein expression in HepG2 cells was abrogated with the suppression of PPARγ by its small interfering RNA or a specific inhibitor, GW9662. These findings suggest that the novel small molecular upregulator 4010B-30 increases ApoA-I gene expression, thereby enhances its function of promoting cholesterol efflux, as well as ABCA1 expression in vitro, and activation of PPARγ is required for 4010B-30 to induce hepatic ApoA-I production. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  2. Folded functional lipid-poor apolipoprotein A-I obtained by heating of high-density lipoproteins: relevance to high-density lipoprotein biogenesis.

    PubMed

    Jayaraman, Shobini; Cavigiolio, Giorgio; Gursky, Olga

    2012-03-15

    HDL (high-density lipoproteins) remove cell cholesterol and protect from atherosclerosis. The major HDL protein is apoA-I (apolipoprotein A-I). Most plasma apoA-I circulates in lipoproteins, yet ~5% forms monomeric lipid-poor/free species. This metabolically active species is a primary cholesterol acceptor and is central to HDL biogenesis. Structural properties of lipid-poor apoA-I are unclear due to difficulties in isolating this transient species. We used thermal denaturation of human HDL to produce lipid-poor apoA-I. Analysis of the isolated lipid-poor fraction showed a protein/lipid weight ratio of 3:1, with apoA-I, PC (phosphatidylcholine) and CE (cholesterol ester) at approximate molar ratios of 1:8:1. Compared with lipid-free apoA-I, lipid-poor apoA-I showed slightly altered secondary structure and aromatic packing, reduced thermodynamic stability, lower self-associating propensity, increased adsorption to phospholipid surface and comparable ability to remodel phospholipids and form reconstituted HDL. Lipid-poor apoA-I can be formed by heating of either plasma or reconstituted HDL. We propose the first structural model of lipid-poor apoA-I which corroborates its distinct biophysical properties and postulates the lipid-induced ordering of the labile C-terminal region. In summary, HDL heating produces folded functional monomolecular lipid-poor apoA-I that is distinct from lipid-free apoA-I. Increased adsorption to phospholipid surface and reduced C-terminal disorder may help direct lipid-poor apoA-I towards HDL biogenesis.

  3. The 5A apolipoprotein A-I mimetic peptide displays anti-inflammatory and antioxidant properties in vivo and in vitro

    PubMed Central

    Tabet, Fatiha; Remaley, Alan T.; Segaliny, Aude I.; Millet, Jonathan; Yan, Ling; Nakhla, Shirley; Barter, Philip J.; Rye, Kerry-Anne; Lambert, Gilles

    2010-01-01

    Objectives The apolipoprotein (apo) A-I mimetic peptide 5A is highly specific for ABCA1-transporter mediated cholesterol efflux. We investigated whether the 5A peptide shares other beneficial features of apoA-I, such as protection against inflammation and oxidation. Methods New-Zealand White rabbits received an infusion of apoA-I, reconstituted HDL containing apoA-I ((A-I)rHDL) or the 5A peptide complexed with phospholipids (PLPC), prior to inserting a collar around the carotid artery. Human coronary artery endothelial cells (HCAECs) were incubated with (A-I)rHDL or 5A/PLPC prior to TNFa stimulation. Results ApoA-I, (A-I)rHDL and 5A/PLPC reduced the collar mediated increase in (i) endothelial expression of cell adhesion molecules VCAM-1 and ICAM-1, (ii) O2− production as well as the expression of the Nox4 catalytic subunits of the NADPH oxidase, and (iii) infiltration of circulating neutrophils into the carotid intima-media. In HCAECs, both 5A/PLPC and (A-I)rHDL inhibited TNFa induced ICAM-1 and VCAM-1 expression as well as the NF-κB signalling cascade and O2− production. The effects of the 5A/PLPC complex were no longer apparent in HCAECs knocked down for ABCA1. Conclusion Like apoA-I, the 5A peptide inhibits acute inflammation and oxidative stress in rabbit carotids and HCAECs. In vitro, the 5A peptide exerts these beneficial effects through interaction with ABCA1. PMID:19965776

  4. Lower HDL-C and apolipoprotein A-I are related to higher glomerular filtration rate in subjects without kidney disease[S

    PubMed Central

    Krikken, Jan A.; Gansevoort, Ron T.; Dullaart, Robin P. F.

    2010-01-01

    Animal experiments show that the kidney contributes to apolipoprotein (apo)A-I catabolism. We tested relationships of HDL cholesterol (HDL-C) and apo-I with kidney function in subjects without severe chronic kidney disease. Included was a random sample of the general population (part of the PREVEND cohort). Kidney function [estimated glomerular filtration rate (e-GFR) by two well-established equations and creatinine clearance], HDL-C, triglycerides, apoA-I and insulin resistance (HOMAir) were measured in 2,484 fasting subjects (e-GFR≥45 ml/min/1.73m2) without macroalbuminuria, cardiovascular disease, diabetes, or the use of anti-hypertensives and/or lipid-lowering agents. HDL-C (r = −0.056 to −0.102, P < 0.01 to < 0.001) and apo A-I (r = −0.096 to −0.126, P < 0.001) were correlated inversely with both GFR estimates and creatinine clearance in univariate analyses. Multiple linear regression analyses also demonstrated inverse relationships of HDL-C and apoA-I with all measures of kidney function even after adjustment for age, sex, waist circumference, HOMAir, triglycerides, and urinary albumin excretion (P = 0.053 to 0.004). In conclusion, HDL-C and apoA-I are inversely related to e-GFR and creatinine clearance in subjects without severely compromised kidney function, which fits the concept that the kidney contributes to apoA-I regulation in humans. High glomerular filtration rate may be an independent determinant of a pro-atherogenic lipoprotein profile. PMID:20211930

  5. Anti-CD20 single chain variable antibody fragment-apolipoprotein A-I chimera containing nanodisks promote targeted bioactive agent delivery to CD20-positive lymphomas.

    PubMed

    Crosby, Natasha M; Ghosh, Mistuni; Su, Betty; Beckstead, Jennifer A; Kamei, Ayako; Simonsen, Jens B; Luo, Bing; Gordon, Leo I; Forte, Trudy M; Ryan, Robert O

    2015-08-01

    A fusion protein comprising an α-CD20 single chain variable fragment (scFv) antibody, a spacer peptide, and human apolipoprotein (apo) A-I was constructed and expressed in Escherichia coli. The lipid interaction properties intrinsic to apoA-I as well as the antigen recognition properties of the scFv were retained by the chimera. scFv•apoA-I was formulated into nanoscale reconstituted high-density lipoprotein particles (termed nanodisks; ND) and incubated with cultured cells. α-CD20 scFv•apoA-I ND bound to CD20-positive non-Hodgkins lymphoma (NHL) cells (Ramos and Granta) but not to CD20-negative T lymphocytes (i.e., Jurkat). Binding to NHL cells was partially inhibited by pre-incubation with rituximab, a monoclonal antibody directed against CD20. Confocal fluorescence microscopy analysis of Granta cells following incubation with α-CD20 scFv•apoA-I ND formulated with the intrinsically fluorescent hydrophobic polyphenol, curcumin, revealed α-CD20 scFv•apoA-I localizes to the cell surface, while curcumin off-loads and gains entry to the cell. Compared to control incubations, viability of cultured NHL cells was decreased upon incubation with α-CD20 scFv•apoA-I ND harboring curcumin. Thus, formulation of curcumin ND with α-CD20 scFv•apoA-I as the scaffold component confers cell targeting and enhanced bioactive agent delivery, providing a strategy to minimize toxicity associated with chemotherapeutic agents.

  6. The expression of intact and mutant human apoAI/CIII/AIV/AV gene cluster in transgenic mice.

    PubMed

    Gao, Jun; Wei, Yusheng; Huang, Yue; Liu, Depei; Liu, Guang; Wu, Min; Wu, Lin; Zhang, Qingjun; Zhang, Zhuqin; Zhang, Ran; Liang, Chihchuan

    2005-04-01

    The apoAI/CIII/AIV gene cluster is involved in lipid metabolism and has a complex pattern of gene expression modulated by a common regulatory element, the apoCIII enhancer. A new member of this cluster, apolipoprotein (apo) AV, has recently been discovered as a novel modifier in triglyceride metabolism. To determine the expression of all four apo genes in combination and, most importantly, whether the transcription of apoAV is coregulated by the apoCIII enhancer in the cluster, we generated an intact transgenic line carrying the 116-kb human apoAI/CIII/AIV/AV gene cluster and a mutant transgenic line in which the apoCIII enhancer was deleted from the 116-kb structure. We demonstrated that the apoCIII enhancer regulated hepatic and intestinal apoAI, apoCIII, and apoAIV expression; however, it did not direct the newly identified apoAV in the cluster. Furthermore, human apo genes displayed integrated position-independent expression and a closer approximation of copy number-dependent expression in the intact transgenic mice. Because apoCIII and apoAV play opposite roles in triglyceride homeostasis, we analyzed the lipid profiles in our transgenic mice to assess the effects of human apoAI gene cluster expression on lipid metabolism. The triglyceride level was elevated in intact transgenic mice but decreased in mutant ones compared with nontransgenic mice. In addition, the expression of human apoAI and apoAIV elevated high density lipoprotein cholesterol in transgenic mice fed an atherogenic diet. In conclusion, our studies with human apoAI/CIII/AIV/AV gene cluster transgenic models showed that the apoCIII enhancer regulated expression of apoAI, apo-CIII, and apoAIV but not apoAV in vivo and showed the influences of expression of the entire cluster on lipid metabolism.

  7. Phylogenetic distribution of apolipoproteins A-I and E in vertebrates as determined by Western blot analysis.

    PubMed

    Duggan, A E; Callard, I P

    2001-08-01

    A putative apolipoprotein E (apoE) has been identified in the HDL and VHDL fractions of the turtle. This observation is of particular interest considering apoE has been reported absent in the domestic hen (Hermier et al., '95; Biochim Biophys Acta: 105-118, 1995) and thus presumed absent in nonmammalian vertebrates altogether. As a result, partial amino acid sequencing of this protein was performed and revealed that one fragment shared 41% sequence identity to human apoE. Western blot analysis using antisera to apoE demonstrated cross-reactivity to a 34-kDa protein (putative apoE) in turtle plasma. Further investigation using anti-apoE antibody in Western blot analysis detected immunoreactive apoE in the plasma of lamprey, spiny dogfish, skate, and alligator, but not in flounder, newt or python; its absence in several species of birds was confirmed. Using anti-apoA-I antibody, apoA-I was detected in all vertebrate groups except a representative teleost (flounder). Apo-A-I antibody cross-reacted weakly with some putative apoE proteins (chicken, spiny dogfish and skate) and the reverse was true for anti-apoE, which cross-reacted with putative apoA-I in birds, reptiles, and elasmobranchs, confirming the molecular similarity and phylogenetic relatedness of these two proteins.

  8. Composition, structure and substrate properties of reconstituted discoidal HDL with apolipoprotein A-I and cholesteryl ester

    NASA Astrophysics Data System (ADS)

    Dergunov, Alexander D.; Shabrova, Elena V.; Dobretsov, Gennady E.

    2010-03-01

    To investigate the influence of lipid unsaturation and neutral lipid on the maturation of high density lipoproteins, the discoidal complexes of apoA-I, phosphatidylcholine and cholesteryl ester (CE) were prepared. Saturated dipalmitoylphosphatidylcholine (DPPC) and unsaturated palmitoyllinoleoylphosphatidylcholine (PLPC), palmitoyloleoylphosphatidylcholine (POPC), and fluorescent probe cholesteryl 1-pyrenedecanoate (CPD) that forms in a diffusion- and concentration-dependent manner short-lived dimer of unexcited and excited molecules (excimer) were used. The apoA-I/DPPC/CPD complexes were heterogeneous by size, composition and probe location. CPD molecules incorporated more efficiently into larger complexes and accumulated in a central part of the discs. The apoA-I/POPC(PLPC)/CPD were also heterogeneous, however, probe molecules distributed preferentially into smaller complexes and accumulated at disc periphery. The kinetics of CPD transfer by recombinant cholesteryl ester transfer protein (CETP) to human plasma LDL is well described by two-exponential decay, the fast component with a shorter transfer time being more populated in PLPC compared to DPPC complexes. The presence of CE molecules in discoidal HDL results in particle heterogeneity. ApoA-I influences the CETP activity modulating the properties of apolipoprotein-phospholipid interface. This may include CE molecules accumulation in the boundary lipid in unsaturated phosphatidylcholine and cluster formation in the bulk bilayer in saturated phosphatidylcholine.

  9. International Federation of Clinical Chemistry standardization project for measurements of apolipoproteins A-I and B. IV. Comparability of apolipoprotein B values by use of International Reference Material.

    PubMed

    Marcovina, S M; Albers, J J; Kennedy, H; Mei, J V; Henderson, L O; Hannon, W H

    1994-04-01

    We performed temporal and thermal stability studies on SP3-07, a liquid-stabilized reference material for apolipoprotein (apo) B, selected during the previous phase of the International Federation of Clinical Chemistry project on standardization of apolipoprotein measurements. Results indicate that SP3-07 stored at -70 degrees C has the long-term stability required for a reference material. We assigned an accuracy-based apo B value of 1.22 g/L to SP3-07, using a nephelometric method that was calibrated with freshly isolated low-density lipoprotein for which the apo B mass value was determined by a standardized sodium dodecyl sulfate-Lowry procedure. Using a common protocol, the study participants transferred the assigned mass value from SP3-07 to the individual calibrators of the analytical systems and measured the apo B concentration of 20 fresh-frozen samples obtained from individual donors and covering a clinically relevant range of apo B values. The among-laboratory CV on these samples, analyzed by 25 analytical systems, ranged from 3.1% to 6.7%. These results demonstrate the lack of matrix effects of SP3-07 and its ability to provide accurate and comparable apo B values in a variety of immunochemical methods. On the basis of the outcome of these studies, the World Health Organization has endorsed SP3-07 as the International Reference Material for Apolipoprotein B.

  10. Serum Apolipoprotein A-I and Large High-Density Lipoprotein Particles Are Positively Correlated with FEV1 in Atopic Asthma

    PubMed Central

    Kaler, Maryann; Cuento, Rosemarie A.; Gordon, Elizabeth M.; Weir, Nargues A.; Sampson, Maureen; Fontana, Joseph R.; MacDonald, Sandra; Moss, Joel; Manganiello, Vincent; Remaley, Alan T.; Levine, Stewart J.

    2015-01-01

    Rationale: Although lipids, apolipoproteins, and lipoprotein particles are important modulators of inflammation, varying relationships exist between these parameters and asthma. Objectives: To determine whether serum lipids and apolipoproteins correlate with the severity of airflow obstruction in subjects with atopy and asthma. Methods: Serum samples were obtained from 154 atopic and nonatopic subjects without asthma, and 159 subjects with atopy and asthma. Serum lipid and lipoprotein levels were quantified using standard diagnostic assays and nuclear magnetic resonance (NMR) spectroscopy. Airflow obstruction was assessed by FEV1% predicted. Measurements and Main Results: Serum lipid levels correlated with FEV1 only in the subjects with atopy and asthma. Serum levels of high-density lipoprotein (HDL) cholesterol and apolipoprotein A-I (apoA-I) were positively correlated with FEV1 in subjects with atopy and asthma, whereas a negative correlation existed between FEV1 and serum levels of triglycerides, low-density lipoprotein (LDL) cholesterol, apolipoprotein B (apoB), and the apoB/apoA-I ratio. NMR spectroscopy identified a positive correlation between FEV1 and HDLNMR particle size, as well as the concentrations of large HDLNMR particles and total IDLNMR (intermediate-density lipoprotein) particles in subjects with atopy and asthma. In contrast, LDLNMR particle size and concentrations of LDLNMR and VLDLNMR (very-low-density lipoprotein) particles were negatively correlated with FEV1 in subjects with atopy and asthma. Conclusions: In subjects with atopy and asthma, serum levels of apoA-I and large HDLNMR particles are positively correlated with FEV1, whereas serum triglycerides, LDL cholesterol, and apoB are associated with more severe airflow obstruction. These results may facilitate future studies to assess whether apoA-I and large HDLNMR particles can reduce airflow obstruction and disease severity in asthma. PMID:25692941

  11. Effect of an isoenergetic traditional Mediterranean diet on apolipoprotein A-I kinetic in men with metabolic syndrome

    PubMed Central

    2013-01-01

    Background The impact of the Mediterranean diet (MedDiet) on high-density lipoprotein (HDL) kinetics has not been studied to date. The objective of this study was therefore to investigate the effect of the MedDiet in the absence of changes in body weight on apolipoprotein (apo) A-I kinetic in men with metabolic syndrome (MetS). Methods Twenty-six men with MetS (NCEP-ATP III) were recruited from the general community. In this fixed sequence study, participants’ diet was first standardized to a control diet reflecting current averages in macronutrient intake in North American men, with all foods and beverages provided under isoenergetic conditions for 5 weeks. Participants were then fed an isoenergetic MedDiet over a subsequent period of 5 weeks to maintain their weight constant. During the last week of each diet, participants received a single bolus dose of [5,5,5-2H3] L-leucine and fasting blood samples were collected at predetermined time points. ApoA-I kinetic was determined by multicompartmental modeling using isotopic enrichment data over time. Data were analyses using MIXED models. Results The response of HDL-cholesterol (C) to MedDiet was heterogeneous, such that there was no mean change compared with the control diet. Plasma apoA-I concentration (−3.9%) and pool size (−5.3%, both P < 0.05) were significantly lower after MedDiet and apoA-I production rate tended to be reduced (−5.7%, P = 0.07) with no change in apoA-I fractional catabolic rate (FCR, -1.6%, P = 0.64). Participants among whom HDL-C concentrations were increased with MedDiet (responders: mean ∆HDL-C: +9.9 ± 3.2%, N = 11) showed significantly greater reductions in apoA-I FCR and in apoB and very-low-density lipoprotein-triglycerides (VLDL-TG) concentrations (all P < 0.04) than those among whom HDL-C levels were reduced after the MedDiet (non-responders: mean ∆HDL-C: -12.0 ± 3.9%, N = 8). Correlation analysis revealed that only variations in apo

  12. Single nucleotide polymorphisms of APOA1 gene and their relationship with serum apolipoprotein A-I concentrations in the native population of Assam

    PubMed Central

    Bora, Kaustubh; Pathak, Mauchumi Saikia; Borah, Probodh; Hussain, Md. Iftikar; Das, Dulmoni

    2015-01-01

    Background There is a growing interest in the role of allelic variants of the APOA1 gene in relation to a number of disorders. We described two common polymorphisms of the APOA1 gene, G-75A and C+83T and investigated their potential influence on the serum apolipoprotein A-I (apo A-I) levels in the native population of Assam — a region that is ethnically distinct and from where no information is hitherto available. Methods Blood samples were collected from 150 healthy volunteers. Apo A-I levels were estimated by immunoturbidometry. Genotyping was done by a PCR-RFLP method that involved DNA extraction from whole blood, followed by polymerase chain reaction and digestion of the PCR product by MspI restriction enzyme, and analysis of fragment sizes in 12% polyacrylamide gel. Results The GG variant at G-75A locus and CC variant at C+83T locus were the most prevalent. GG/CC was the most common combination. Homozygous TT genotype was not detected in any of the subjects. The rare allele frequencies for the G-75A and C+83T sites were found to be 0.22 and 0.06 respectively, which significantly differed from those reported in some other populations in neighbouring regions. Serum apo A-I concentrations did not vary significantly across the detected genotypes. These findings were consistent in both sexes. Conclusion We described the distribution of the G-75A and C+83T polymorphisms of the APOA1 gene in the population of Assam for the first time. These polymorphisms were not found to directly influence apo A-I concentrations in this population either individually or synergistically. PMID:26702398

  13. Immunochemical determination of human apolipoprotein B by laser nephelometry.

    PubMed

    Fievet-Desreumaux, C; Dedonder-Decoopman, E; Fruchart, J C; Dewailly, P; Sezille, G

    1979-07-16

    The Hyland laser nephelometer PDQ system for the assay of apolipoprotein B (apo-B) in human serum is described. Within and between-batch precision, accuracy and reliability are discussed. This instrument represents an important development in the immunochemical assay of apo-B, and the speed, precision, and convenience of the methodology make such a system attractive. Quantitation of apo-B was assessed in normal and hyperlipaemic subjects. Comparisons were made with two other specific and sensitive immunological methods for quantifying apo-B: enzymeimmunoassay (EIA) and rocket immunoelectrophoresis (RIE). Results obtained by the three methods correlated very well.

  14. Cholesterol can stimulate secretion of apolipoprotein B by cultured human hepatocytes.

    PubMed

    Kosykh, V A; Preobrazhensky, S N; Fuki, I V; Zaikina, O E; Tsibulsky, V P; Repin, V S; Smirnov, V N

    1985-10-02

    During a 5 day cultivation of human hepatocytes in a primary culture the secretion of apolipoprotein B was measured by enzyme-linked immunosorbent assay. Density-gradient ultracentrifugation demonstrated that the majority of the secreted apolipoprotein B was associated with the very-low-density lipoprotein fraction. Exposure of the cells to cholesterol (5-100 micrograms/ml) resulted in a dose-dependent increase in apolipoprotein B secretion rate.

  15. The intrinsic factor-vitamin B12 receptor, cubilin, is a high-affinity apolipoprotein A-I receptor facilitating endocytosis of high-density lipoprotein.

    PubMed

    Kozyraki, R; Fyfe, J; Kristiansen, M; Gerdes, C; Jacobsen, C; Cui, S; Christensen, E I; Aminoff, M; de la Chapelle, A; Krahe, R; Verroust, P J; Moestrup, S K

    1999-06-01

    Cubilin is the intestinal receptor for the endocytosis of intrinsic factor-vitamin B12. However, several lines of evidence, including a high expression in kidney and yolk sac, indicate it may have additional functions. We isolated apolipoprotein A-I (apoA-I), the main protein of high-density lipoprotein (HDL), using cubilin affinity chromatography. Surface plasmon resonance analysis demonstrated a high-affinity binding of apoA-I and HDL to cubilin, and cubilin-expressing yolk sac cells showed efficient 125I-HDL endocytosis that could be inhibited by IgG antibodies against apoA-I and cubilin. The physiological relevance of the cubilin-apoA-I interaction was further emphasized by urinary apoA-I loss in some known cases of functional cubilin deficiency. Therefore, cubilin is a receptor in epithelial apoA-I/HDL metabolism.

  16. Pig major acute-phase protein and apolipoprotein A-I responses correlate with the clinical course of experimentally induced African Swine Fever and Aujeszky's disease.

    PubMed

    Carpintero, Rakel; Alonso, Covadonga; Piñeiro, Matilde; Iturralde, María; Andrés, Marta; Le Potier, Marie-Frédérique; Madec, Francois; Alava, María A; Piñeiro, Andrés; Lampreave, Fermín

    2007-01-01

    In the present work, we studied the acute phase protein response after experimental virus infection in pigs. The animals were experimentally infected with African Swine Fever (ASF) or Aujeszky's disease (AD) viruses. The clinical course of ASF infection correlated with increasingly high levels of pig Major Acute-phase Protein (pig-MAP) (mean value of 6 mg/mL on day 6 post infection (p.i.), from 6 to 9 times higher than day 0) and sharp apolipoprotein A-I (apo A-I) decrease (mean value of 0.5 mg/mL, from 4 to 10 times lower than day 0 on day 4 p.i.). AD-clinical signs appeared at day 3 p.i., both in vaccinated (moderate clinical signs) and non-vaccinated pigs (severe outcome within 48 h p.i.). Pig-MAP and apo A-I profiles also followed clinical signs (changing from 0.70 mg/mL to around 3 mg/mL and from around 3 mg/mL to 0.96 mg/mL, respectively in non-vaccinated animals), with minor changes in concentration in the vaccinated group. Haptoglobin levels significantly increased in ASF and AD infected animals (mean maximum values of 2.77 and 3.96 mg/mL, respectively). Minor differences for the C-Reactive Protein in the case of ASF were observed, whereas its concentration increased more than 7 times in AD-infection. The albumin level was not modified in either case. The correlation of clinical signs to our data suggests the potential use of pig-MAP and apo A-I in monitoring infections in swine.

  17. Concomitant Effects of Ramadan Fasting and Time-Of-Day on Apolipoprotein AI, B, Lp-a and Homocysteine Responses during Aerobic Exercise in Tunisian Soccer Players

    PubMed Central

    Hammouda, Omar; Chtourou, Hamdi; Aloui, Asma; Chahed, Henda; Kallel, Choumous; Miled, Abdelhedi; Chamari, Karim; Chaouachi, Anis; Souissi, Nizar

    2013-01-01

    Objective To examine the time-of-day and Ramadan fasting (RF) effects on serum apolipoprotein-AI (Apo-AI) and B (Apo-B), lipoprotein particles-a (Lp-a), high-sensitive C-reactive-protein (hs-CRP), and homocysteine (Hcy) during the Yo-Yo intermittent recovery test (YYIRT). Design Performance and biochemical measures were completed at two times-of-day (07:00 and 17:00 h), 1-week before RF (BR), the second week of RF (SWR), and the fourth week of RF (ER). Setting For each session, subjects performed the YYIRT, and blood samples were taken before and 3-min after the test for biochemical measures. Participants Fifteen soccer players. Main Outcome Measures Total distance during the YYIRT, core temperature, body composition, dietary intakes, lipid (HDL-C, LDL-C, Apo-AI, B and Lp-a) and inflammatory (hs-CRP and Hcy) profiles. Results Performances during the YYIRT were higher in the evening than the morning BR (P < 0.05), but this fluctuation was not observed during RF. Moreover, LDL-C, ApoB, and Lp-a were stable throughout the daytime BR. However, during RF, they decreased at 17:00 h (P < 0.05). Likewise, HDL-C and Apo-AI increased after the exercise and were higher at 17:00 h BR (P < 0.001). Moreover, these parameters increased during RF (P < 0.01). Furthermore, Hcy and hs-CRP increased during the exercise (P < 0.01) with higher evening levels BR. During ER, the diurnal pattern of Hcy was inversed (P < 0.001). Conclusions This study concluded that caloric restriction induced by RF seems to ameliorate lipid and inflammatory markers of cardiovascular health during intermittent exercise performed in the evening. PMID:24244572

  18. Corneal vesicles accumulate collagen VI associated with tissue remodeling in apolipoprotein a-I deficiency: a case report.

    PubMed

    Namba, Hiroyuki; Narumi, Mari; Susa, Shinji; Ohe, Rintaro; Kato, Takeo; Yamakawa, Mitsunori; Yamashita, Hidetoshi

    2017-02-08

    Apo A-I deficiency clinically shows low serum levels of HDL cholesterol and corneal opacity at a young age. Histopathological evaluations of affected corneas are not enough, and the mechanism of corneal opacity is still unclear. A 61-year-old woman suffered from blurred vision with a corneal opacity. She had significantly reduced serum levels of high-density lipoprotein cholesterol and Apo A-I, stenosis of the coronary arteries, and ischemic heart failure. On genetic examination, a homozygous mutation of Apo A-ITsukuba was identified. Histopathological examination of the corneal button after PKP showed numerous vesicles in the corneal stroma, which were more prominent in the deep stroma than in the shallow stroma. Collagen VI was observed in some of those vesicles. We experienced a rare case of corneal opacity due to Apo A-I deficiency. Our histopathological findings indicated that structural changes in corneal collagen fibrils contribute to the formation of stromal vesicles.

  19. A human apolipoprotein E mimetic peptide reduces atherosclerosis in aged apolipoprotein E null mice

    PubMed Central

    Xu, Yanyong; Liu, Hongmei; Liu, Mengting; Li, Feifei; Liu, Liangchen; Du, Fen; Fan, Daping; Yu, Hong

    2016-01-01

    Apolipoprotein E (apoE) is well known as an antiatherogenic protein via regulating lipid metabolism and inflammation. We previously reported that a human apoE mimetic peptide, EpK, reduced atherosclerosis in apoE null (apoE-/-) mice through reducing inflammation without affecting plasma lipid levels. Here, we construct another human apoE mimetic peptide, named hEp, and investigate whether expression of hEp can reduce atherosclerotic lesion development in aged female apoE-/- mice with pre-existing lesions. We found that chemically synthesized hEp significantly decreased cholesterol accumulation induced by oxidized low density lipoprotein and the expression of inflammatory cytokines TNFα and IL-6 induced by lipopolysaccharide in macrophages. In an in vivo study, Lv-hEp-GFP lentiviruses were intravenously injected into 9 month-old apoE-/- mice. Mice were then fed a chow diet for 18 weeks. Results showed that in comparison to the Lv-GFP lentivirus injection (Lv-GFP) group, Lv-hEp-GFP lentivirus injection achieved hepatic hEp expression and secretion in apoE-/- mice. It was observed that hEp expression significantly reduced plasma VLDL and LDL cholesterol levels and decreased aortic atherosclerotic lesions. This was accompanied by an increase of LDL receptor expression and a reduction of TNFα and IL-6 mRNA levels in the liver. Moreover, expression of hEp increased plasma paraoxonase-1 activity and decreased plasma myeloperoxidase activity and serum amyloid A levels. Our study provides evidence that hEp may be developed as a promising therapeutic apoE mimetic peptide for atherosclerosis-related cardiovascular diseases through its induction of plasma VLDL/LDL cholesterol clearance as well as its anti-oxidative and anti-inflammatory activities. PMID:27648138

  20. A novel mutation of the apolipoprotein A-I gene in a family with familial combined hyperlipidemia.

    PubMed

    Pisciotta, Livia; Fasano, Tommaso; Calabresi, Laura; Bellocchio, Antonella; Fresa, Raffaele; Borrini, Claudia; Calandra, Sebastiano; Bertolini, Stefano

    2008-05-01

    We report a large family in which four members showed a plasma lipid profile consistent with the clinical diagnosis of familial combined hyperlipidemia (FCHL). One of these patients was found to have markedly reduced HDL cholesterol (HDL-C) (0.72 mmol/l) and Apo A-I (72 mg/dl) levels, a condition suggestive of the presence of a mutation in one of the HDL-related genes. The analysis of APOA1 gene revealed that this patient was heterozygous for a cytosine insertion in exon 3 (c.49-50 ins C), resulting in a frame-shift and premature stop codon at position 26 of pro-Apo A-I (Q17PFsX10). This novel mutation, which prevents the synthesis of Apo A-I, was also found in four family members, including three siblings and the daughter of the proband. Carriers of Apo A-I mutation had significantly lower HDL-C and Apo A-I than non-carriers family members (0.77+/-0.15 mmol/l vs. 1.15+/-0.20 mmol/l, P<0.005; 71.4+/-9.1mg/dl vs. 134.0+/-14.7 mg/dl, P<0.005, respectively). Two of the APOA1 mutation carriers, who were also heavy smokers, had fibrous plaques in the carotid arteries causing mild stenosis (20%). The intimal-media thickness in the two other adult carriers was within the normal range. The other non-carriers family members with FCHL had either overt vascular disease or carotid atherosclerosis at ultrasound examination. This observation suggests that the low HDL-C/low Apo A-I phenotype may result from a genetic defect directly affecting HDL metabolism, even in the context of a dyslipidemia which, like FCHL, is associated with low plasma HDL-C.

  1. Introduction of human apolipoprotein E4 "domain interaction" into mouse apolipoprotein E.

    PubMed

    Raffai, R L; Dong, L M; Farese, R V; Weisgraber, K H

    2001-09-25

    Human apolipoprotein E4 (apoE4) binds preferentially to lower density lipoproteins, including very low density lipoproteins, and is associated with increased risk of atherosclerosis and neurodegenerative disorders, including Alzheimer's disease. This binding preference is the result of the presence of Arg-112, which causes Arg-61 in the amino-terminal domain to interact with Glu-255 in the carboxyl-terminal domain. ApoE2 and apoE3, which have Cys-112, bind preferentially to high density lipoproteins (HDL) and do not display apoE4 domain interaction. Mouse apoE, like apoE4, contains the equivalent of Arg-112 and Glu-255, but lacks the critical Arg-61 equivalent (it contains Thr-61). Thus, mouse apoE does not display apoE4 domain interaction and, as a result, behaves like human apoE3, including preferential binding to HDL. To assess the potential role of apoE4 domain interaction in atherosclerosis and neurodegeneration, we sought to introduce apoE4 domain interaction into mouse apoE. Replacing Thr-61 in mouse apoE with arginine converted the binding preference from HDL to very low density lipoproteins in vitro, suggesting that apoE4 domain interaction could be introduced into mouse apoE in vivo. Using gene targeting in embryonic stem cells, we created mice expressing Arg-61 apoE. Heterozygous Arg-61/wild-type apoE mice displayed two phenotypes found in human apoE4/E3 heterozygotes: preferential binding to lower density lipoproteins and reduced abundance of Arg-61 apoE in the plasma, reflecting its more rapid catabolism. These findings demonstrate the successful introduction of apoE4 domain interaction into mouse apoE in vivo. The Arg-61 apoE mouse model will allow the effects of apoE4 domain interaction in lipoprotein metabolism, atherosclerosis, and neurodegeneration to be determined.

  2. Human apolipoprotein E expression in Escherichia coli: structural and functional identity of the bacterially produced protein with plasma apolipoprotein E.

    PubMed Central

    Vogel, T; Weisgraber, K H; Zeevi, M I; Ben-Artzi, H; Levanon, A Z; Rall, S C; Innerarity, T L; Hui, D Y; Taylor, J M; Kanner, D

    1985-01-01

    Human apolipoprotein E (apoE) was produced in Escherichia coli by transforming cells with an expression vector containing a reconstructed apoE cDNA, a lambda PL promoter regulated by the thermolabile cI repressor, and a ribosomal binding site derived from the lambda cII or the E. coli beta-lactamase gene. Transformed cells induced at 42 degrees C for short periods of time (less than 20 min) produced apoE, which accumulated in the cells at levels of approximately equal to 1% of the total soluble cellular protein. Longer induction periods resulted in cell lysis and the proteolytic destruction of apoE. The bacterially produced apoE was purified by heparin-Sepharose affinity chromatography, Sephacryl S-300 gel filtration, and preparative Immobiline isoelectric focusing. The final yield was approximately equal to 20% of the initial apoE present in the cells. Except for an additional methionine at the amino terminus, the bacterially produced apoE was indistinguishable from authentic human plasma apoE as determined by NaDodSO4 and isoelectric focusing gel electrophoresis, amino acid composition of the total protein as well as its cyanogen bromide fragments, and partial amino acid sequence analysis (residues 1-17 and 109-164). Both the bacterially produced and authentic plasma apoE bound similarly to apolipoprotein B,E(low density lipoprotein) receptors of human fibroblasts and to hepatic apoE receptors. Intravenous injection resulted in similar rates of clearance for both the bacterially produced and authentic apoE from rabbit and rat plasma (approximately equal to 50% removed in 20 min). The ability to synthesize a bacterially produced human apolipoprotein with biological properties indistinguishable from those of the native protein will allow the production of large quantities of apoE for use in further investigations of the biological and physiological properties of this apolipoprotein. Images PMID:3909150

  3. TRL, IDL, and LDL apolipoprotein B-100 and HDL apolipoprotein A-I kinetics as a function of age and menopausal status.

    PubMed

    Matthan, Nirupa R; Jalbert, Susan M; Lamon-Fava, Stefania; Dolnikowski, Gregory G; Welty, Francine K; Barrett, Hugh R; Schaefer, Ernst J; Lichtenstein, Alice H

    2005-08-01

    To determine mechanisms contributing to the altered lipoprotein profile associated with aging and menopause, apolipoprotein B-100 (apoB-100) and apoA-I kinetic behavior was assessed. Eight premenopausal (25+/-3 years) and 16 postmenopausal (65+/-6 years) women consumed for 6 weeks a standardized Western diet, at the end of which a primed-constant infusion of deuterated leucine was administered in the fed state to determine the kinetic behavior of triglyceride-rich lipoprotein (TRL), intermediate-density lipoprotein (IDL), and low-density lipoprotein (LDL) apoB-100, and high-density lipoprotein (HDL) apoA-I. Data were fit to a multicompartmental model using SAAM II to calculate fractional catabolic rate (FCR) and production rate (PR). Total cholesterol, LDL cholesterol (LDL-C), TRL-C, and triglyceride levels were higher (50%, 55%, 130%, and 232%, respectively) in the postmenopausal compared with the premenopausal women, whereas HDL-C levels were similar. Plasma TRL, IDL, and LDL-apoB-100 levels and pool sizes (PS) were significantly higher in the postmenopausal than premenopausal women. These differences were accounted for by lower TRL, IDL, and LDL apoB-100 FCR (P<0.05), with no difference in PR. There was no significant difference between groups in HDL-C levels or apoA-I kinetic parameters. Plasma TRL-C concentrations were negatively correlated with TRL apoB-100 FCR (r=-0.46; P<0.05) and positively correlated with PR (r=0.62; P<0.01). Plasma LDL-C concentrations were negatively correlated with LDL apoB-100 FCR (r=-0.70; P<0.001) but not PR. The mechanism for the increase in TRL and LDL apoB-100 PS observed in the postmenopausal women was determined predominantly by decreased TRL and LDL catabolism rather than increased production. No differences were observed in HDL apoA-I kinetics between groups.

  4. Regulation of the human apolipoprotein AIV gene expression in transgenic mice.

    PubMed

    Baralle, M; Vergnes, L; Muro, A F; Zakin, M M; Baralle, F E; Ochoa, A

    1999-02-19

    The apolipoprotein (Apo) AI-CIII-AIV gene cluster has a complex pattern of gene expression that is modulated by both gene- and cluster-specific cis-acting elements. In particular the regulation of Apo AIV expression has been previously studied in vivo and in vitro including several transgenic mouse lines but a complete, consistent picture of the tissue-specific controls is still missing. We have analysed the role of the Apo AIV 3' flanking sequences in the regulation of gene expression using both in vitro and in vivo systems including three lines of transgenic mice. The transgene consisted of a human fragment containing 7 kb of the 5' flanking region, the Apo AIV gene itself and 6 kb of the 3' flanking region (-7+6 Apo AIV). Accurate analysis of the Apo AIV mRNA levels using quantitative PCR and Northern blots showed that the 7+6 kb Apo AIV fragment confers liver-specific regulation in that the human Apo AIV transgene is expressed at approximately the same level as the endogenous mouse Apo AIV gene. In contrast, the intestinal regulation of the transgene did not follow, the pattern observed with the endogenous gene although it produced a much higher intestinal expression following the accepted human pattern. Therefore, this animal model provides an excellent substrate to design therapeutic protocols for those metabolic derangements that may benefit from variations in Apo AIV levels and its anti-atherogenic effect.

  5. Diabetic foot disease: grading inflammation by apolipoprotein A-I, C-reactive protein and serum amyloid A.

    PubMed

    Wang, Weiling; Zhang, Yan; Liao, Yonggan; Wu, Jun; Zhou, Lixia; Tang, Yijun; Cao, Yang; Metzmann, Erwin; Wang, Rongfang

    2014-01-01

    The different grading systems for diabetic foot disease pose a challenge in clinical decision making because each system fails to accurately reflect the individual course of disease progression. This study attempts to identify laboratory measurements for classifying diabetic foot disease to guide clinical treatment. The sera of 111 clinically graded diabetic foot patients were measured for several laboratory parameters including serum amyloid A (SAA), C-reactive protein (CRP), and apo A-I. By using the molar sum of CRP and SAA and then dividing by the molarity of apo A-I, an acute phase index was introduced to assess the inflammatory status of the patients. Based on a newly defined acute phase index (API), diabetic foot patients were classified into 3 distinct groups that provide a diagnostic tool complementing the established Texas grading system for clinical decision making. The integration of the serum concentrations of SAA, CRP and apo A-I into an acute phase index (API) offers an opportunity to triage diabetic foot patients who may benefit from personalized medicine.

  6. Levels and changes of HDL cholesterol and apolipoprotein A-I in relation to risk of cardiovascular events among statin-treated patients: a meta-analysis.

    PubMed

    Boekholdt, S Matthijs; Arsenault, Benoit J; Hovingh, G Kees; Mora, Samia; Pedersen, Terje R; Larosa, John C; Welch, K M A; Amarenco, Pierre; Demicco, David A; Tonkin, Andrew M; Sullivan, David R; Kirby, Adrienne; Colhoun, Helen M; Hitman, Graham A; Betteridge, D John; Durrington, Paul N; Clearfield, Michael B; Downs, John R; Gotto, Antonio M; Ridker, Paul M; Kastelein, John J P

    2013-10-01

    It is unclear whether levels of high-density lipoprotein cholesterol (HDL-C) or apolipoprotein A-I (apoA-I) remain inversely associated with cardiovascular risk among patients who achieve very low levels of low-density lipoprotein cholesterol on statin therapy. It is also unknown whether a rise in HDL-C or apoA-I after initiation of statin therapy is associated with a reduced cardiovascular risk. We performed a meta-analysis of 8 statin trials in which lipids and apolipoproteins were determined in all study participants at baseline and at 1-year follow-up. Individual patient data were obtained for 38,153 trial participants allocated to statin therapy, of whom 5387 suffered a major cardiovascular event. HDL-C levels were associated with a reduced risk of major cardiovascular events (adjusted hazard ratio [HR], 0.83; 95% confidence interval [CI], 0.81-0.86 per 1 standard deviation increment), as were apoA-I levels (HR, 0.79; 95% CI, 0.72-0.82). This association was also observed among patients achieving on-statin low-density lipoprotein cholesterol levels <50 mg/dL. An increase of HDL-C was not associated with reduced cardiovascular risk (HR, 0.98; 95% CI, 0.94-1.01 per 1 standard deviation increment), whereas a rise in apoA-I was (HR, 0.93; 95% CI, 0.90-0.97). Among patients treated with statin therapy, HDL-C and apoA-I levels were strongly associated with a reduced cardiovascular risk, even among those achieving very low low-density lipoprotein cholesterol. An apoA-I increase was associated with a reduced risk of major cardiovascular events, whereas for HDL-C this was not the case. These findings suggest that therapies that increase apoA-I concentration require further exploration with regard to cardiovascular risk reduction.

  7. Interaction of an atypical Plasmodium falciparum ETRAMP with human apolipoproteins

    PubMed Central

    Vignali, Marissa; McKinlay, Anastasia; LaCount, Douglas J; Chettier, Rakesh; Bell, Russell; Sahasrabudhe, Sudhir; Hughes, Robert E; Fields, Stanley

    2008-01-01

    Background In order to establish a successful infection in the human host, the malaria parasite Plasmodium falciparum must establish interactions with a variety of human proteins on the surface of different cell types, as well as with proteins inside the host cells. To better understand this aspect of malaria pathogenesis, a study was conducted with the goal of identifying interactions between proteins of the parasite and those of its human host. Methods A modified yeast two-hybrid methodology that preferentially selects protein fragments that can be expressed in yeast was used to conduct high-throughput screens with P. falciparum protein fragments against human liver and cerebellum libraries. The resulting dataset was analyzed to exclude interactions that are not likely to occur in the human host during infection. Results An initial set of 2,200 interactions was curated to remove proteins that are unlikely to play a role in pathogenesis based on their annotation or localization, and proteins that behave promiscuously in the two-hybrid assay, resulting in a final dataset of 456 interactions. A cluster that implicates binding between P. falciparum PFE1590w/ETRAMP5, a putative parasitophorous vacuole membrane protein, and human apolipoproteins ApoA, ApoB and ApoE was selected for further analysis. Different isoforms of ApoE, which are associated with different outcomes of malaria infection, were shown to display differential interactions with PFE1590w. Conclusion A dataset of interactions between proteins of P. falciparum and those of its human host was generated. The preferential interaction of the P. falciparum PFE1590w protein with the human ApoE ε3 and ApoE ε4 isoforms, but not the ApoE ε2 isoform, supports the hypothesis that ApoE genotype affects risk of malaria infection. The dataset contains other interactions of potential relevance to disease that may identify possible vaccine candidates and drug targets. PMID:18937849

  8. Selective HDL-Raising Human Apo A-I Gene Therapy Counteracts Cardiac Hypertrophy, Reduces Myocardial Fibrosis, and Improves Cardiac Function in Mice with Chronic Pressure Overload.

    PubMed

    Amin, Ruhul; Muthuramu, Ilayaraja; Aboumsallem, Joseph Pierre; Mishra, Mudit; Jacobs, Frank; De Geest, Bart

    2017-09-20

    Epidemiological studies support an independent inverse association between high-density lipoprotein (HDL) cholesterol levels and heart failure incidence. The effect of selective HDL-raising adeno-associated viral serotype 8-human apolipoprotein (apo) A-I (AAV8-A-I) gene transfer on cardiac remodeling induced by transverse aortic constriction (TAC) was evaluated in C57BL/6 low-density lipoprotein receptor-deficient mice. Septal wall thickness and cardiomyocyte cross-sectional area were reduced by 16.5% (p < 0.001) and by 13.8% (p < 0.01), respectively, eight weeks after TAC in AAV8-A-I mice (n = 24) compared to control mice (n = 39). Myocardial capillary density was 1.11-fold (p < 0.05) higher and interstitial cardiac fibrosis was 45.3% (p < 0.001) lower in AAV8-A-I TAC mice than in control TAC mice. Lung weight and atrial weight were significantly increased in control TAC mice compared to control sham mice, but were not increased in AAV8-A-I TAC mice. The peak rate of isovolumetric contraction was 1.19-fold (p < 0.01) higher in AAV8-A-I TAC mice (n = 17) than in control TAC mice (n = 29). Diastolic function was also significantly enhanced in AAV8-A-I TAC mice compared to control TAC mice. Nitro-oxidative stress and apoptosis were significantly reduced in the myocardium of AAV8-A-I TAC mice compared to control TAC mice. In conclusion, selective HDL-raising human apo A-I gene transfer potently counteracts the development of pressure overload-induced cardiomyopathy.

  9. Apolipoprotein E promotes lipid accumulation and differentiation in human adipocytes

    SciTech Connect

    Lasrich, Dorothee; Bartelt, Alexander; Grewal, Thomas; Heeren, Joerg

    2015-09-10

    Several studies in mice indicate a role for apolipoprotein E (APOE) in lipid accumulation and adipogenic differentiation in adipose tissue. However, little is yet known if APOE functions in a similar manner in human adipocytes. This prompted us to compare lipid loading and expression of adipocyte differentiation markers in APOE-deficient and control adipocytes using the differentiated human mesenchymal stem cell line hMSC-Tert as well as primary human and mouse adipocytes as model systems. Differentiated hMSC-Tert were stably transduced with or without siRNA targeting APOE while murine adipocytes were isolated from wild type and Apoe knockout mice. Human APOE knockdown hMSC-Tert adipocytes accumulated markedly less triglycerides compared to control cells. This correlated with strongly decreased gene expression levels of adipocyte markers such as adiponectin (ADIPOQ) and fatty acid binding protein 4 (FABP4) as well as the key transcription factor driving adipocyte differentiation, peroxisome proliferator activator receptor gamma (PPARG), in particular the PPARG2 isoform. Similarly, differentiation of murine Apoe-deficient adipocytes was characterized by reduced gene expression of Adipoq, Fabp4 and Pparg. Interestingly, incubation of APOE-deficient hMSC-Tert adipocytes with conditioned media from APOE3-overexpressing adipocytes or APOE-containing Very Low Density Lipoprotein (VLDL) partially restored triglyceride accumulation, but were unable to induce adipocyte differentiation, as judged by expression of adipocyte markers. Taken together, depletion of endogenous APOE in human adipocytes severely impairs lipid accumulation, which is associated with an inability to initiate differentiation. - Highlights: • Immortalized human mesenchymal stem cells were used to study adipocyte development. • Knockdown of endogenous APOE lead to impaired lipid accumulation and adipogenesis. • APOE supplementation partially restored lipid accumulation but not differentiation.

  10. An Evaluation of the Crystal Structure of C-terminal Truncated Apolipoprotein A-I in Solution Reveals Structural Dynamics Related to Lipid Binding.

    PubMed

    Melchior, John T; Walker, Ryan G; Morris, Jamie; Jones, Martin K; Segrest, Jere P; Lima, Diogo B; Carvalho, Paulo C; Gozzo, Fábio C; Castleberry, Mark; Thompson, Thomas B; Davidson, W Sean

    2016-03-04

    Apolipoprotein (apo) A-I mediates many of the anti-atherogenic functions attributed to high density lipoprotein. Unfortunately, efforts toward a high resolution structure of full-length apoA-I have not been fruitful, although there have been successes with deletion mutants. Recently, a C-terminal truncation (apoA-I(Δ185-243)) was crystallized as a dimer. The structure showed two helical bundles connected by a long, curved pair of swapped helical domains. To compare this structure to that existing under solution conditions, we applied small angle x-ray scattering and isotope-assisted chemical cross-linking to apoA-I(Δ185-243) in its dimeric and monomeric forms. For the dimer, we found evidence for the shared domains and aspects of the N-terminal bundles, but not the molecular curvature seen in the crystal. We also found that the N-terminal bundles equilibrate between open and closed states. Interestingly, this movement is one of the transitions proposed during lipid binding. The monomer was consistent with a model in which the long shared helix doubles back onto the helical bundle. Combined with the crystal structure, these data offer an important starting point to understand the molecular details of high density lipoprotein biogenesis. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Cholesterol Efflux Capacity of Apolipoprotein A-I Varies with the Extent of Differentiation and Foam Cell Formation of THP-1 Cells

    PubMed Central

    Yano, Kouji; Sato, Megumi; Yoshimoto, Akira; Ichimura, Naoya; Kameda, Takahiro; Kubota, Tetsuo

    2016-01-01

    Apolipoprotein A-I (apoA-I), the main protein component of high-density lipoprotein (HDL), has many protective functions against atherosclerosis, one of them being cholesterol efflux capacity. Although cholesterol efflux capacity measurement is suggested to be a key biomarker for evaluating the risk of development of atherosclerosis, the assay has not been optimized till date. This study aims at investigating the effect of different states of cells on the cholesterol efflux capacity. We also studied the effect of apoA-I modification by homocysteine, a risk factor for atherosclerosis, on cholesterol efflux capacity in different states of cells. The cholesterol efflux capacity of apoA-I was greatly influenced by the extent of differentiation of THP-1 cells and attenuated by excessive foam cell formation. N-Homocysteinylated apoA-I indicated a lower cholesterol efflux capacity than normal apoA-I in the optimized condition, whereas no significant difference was observed in the cholesterol efflux capacity between apoA-I in the excessive cell differentiation or foam cell formation states. These results suggest that cholesterol efflux capacity of apoA-I varies depending on the state of cells. Therefore, the cholesterol efflux assay should be performed using protocols optimized according to the objective of the experiment. PMID:27957343

  12. Serum amyloid A protein forms a complex with a fragment of apolipoprotein A-I in the domestic blue fox: a protective mechanism against AA amyloidosis?

    PubMed

    Elisen, Ellen Johanne; Bruun, Cathrine Foyn; Nordstoga, Knut; Husby, Gunnar; Sletten, Knut

    2004-09-01

    The spontaneous occurrence of protein AA-type of amyloidosis varies among animal species. As reactive AA-type of amyloidosis has never been detected in the blue fox, we obtained acute phase sera to search for amyloid-protective elements. The purified SAA fraction was characterized by mass and sequence analyses to disclose any unique domains in the amino acid sequence. The data revealed an SAA protein with heterogeneities in several positions, and showed the typical insertion between positions 69 and 70. By comparing the amino acid sequence with that from other mammals, no unique sequence could be observed. However, a C-terminal fragment of apolipoprotein A-I (ApoA-I) was found attached to the SAA. The amino acid sequence of the ApoA-I fragment revealed a partially blocked and ragged N-terminus. A comparison of the amino acid sequence of ApoA-I with that from the dog showed that the ApoA-I fragment started about position 190, had an intact C-terminus, and showed an identical sequence in all positions, except one. Based on the data, we suggest an interaction of the C-terminal fragment of ApoA-I with the SAA protein that inhibits the AA fibrillogenesis in the blue fox.

  13. An apolipoprotein A-I mimetic peptide designed with a reductionist approach stimulates reverse cholesterol transport and reduces atherosclerosis in mice.

    PubMed

    Ditiatkovski, Michael; D'Souza, Wilissa; Kesani, Rajitha; Chin-Dusting, Jaye; de Haan, Judy B; Remaley, Alan; Sviridov, Dmitri

    2013-01-01

    Apolipoprotein A-I (apoA-I) mimetic peptides are considered a promising novel therapeutic approach to prevent and/or treat atherosclerosis. An apoA-I mimetic peptide ELK-2A2K2E was designed with a reductionist approach and has shown exceptional activity in supporting cholesterol efflux but modest anti-inflammatory and anti-oxidant properties in vitro. In this study we compared these in vitro properties with the capacity of this peptide to modify rates of reverse cholesterol transport and development of atherosclerosis in mouse models. The peptide enhanced the rate of reverse cholesterol transport in C57BL/6 mice and reduced atherosclerosis in Apoe(-/-) mice receiving a high fat diet. The peptide modestly reduced the size of the plaques in aortic arch, but was highly active in reducing vascular inflammation and oxidation. Administration of the peptide to Apoe(-/-) mice on a high fat diet reduced the levels of total, high density lipoprotein and non-high density lipoprotein cholesterol and triglycerides. It increased the proportion of smaller HDL particles in plasma at the expense of larger HDL particles, and increased the capacity of the plasma to support cholesterol efflux. Thus, ELK-2A2K2E peptide reduced atherosclerosis in Apoe(-/-) mice, however, the functional activity profile after chronic in vivo administration was different from that found in acute in vitro studies.

  14. Tamarindus indica extract alters release of alpha enolase, apolipoprotein A-I, transthyretin and Rab GDP dissociation inhibitor beta from HepG2 cells.

    PubMed

    Chong, Ursula Rho Wan; Abdul-Rahman, Puteri Shafinaz; Abdul-Aziz, Azlina; Hashim, Onn Haji; Junit, Sarni Mat

    2012-01-01

    The plasma cholesterol and triacylglycerol lowering effects of Tamarindus indica extract have been previously described. We have also shown that the methanol extract of T. indica fruit pulp altered the expression of lipid-associated genes including ABCG5 and APOAI in HepG2 cells. In the present study, effects of the same extract on the release of proteins from the cells were investigated using the proteomics approach. When culture media of HepG2 cells grown in the absence and presence of the methanol extract of T. indica fruit pulp were subjected to 2-dimensional gel electrophoresis, the expression of seven proteins was found to be significantly different (p<0.03125). Five of the spots were subsequently identified as alpha enolase (ENO1), transthyretin (TTR), apolipoprotein A-I (ApoA-I; two isoforms), and rab GDP dissociation inhibitor beta (GDI-2). A functional network of lipid metabolism, molecular transport and small molecule biochemistry that interconnects the three latter proteins with the interactomes was identified using the Ingenuity Pathways Analysis software. The methanol extract of T. indica fruit pulp altered the release of ENO1, ApoA-I, TTR and GDI-2 from HepG2 cells. Our results provide support on the effect of T. indica extract on cellular lipid metabolism, particularly that of cholesterol.

  15. An Evaluation of the Crystal Structure of C-terminal Truncated Apolipoprotein A-I in Solution Reveals Structural Dynamics Related to Lipid Binding*

    PubMed Central

    Melchior, John T.; Walker, Ryan G.; Morris, Jamie; Jones, Martin K.; Segrest, Jere P.; Lima, Diogo B.; Carvalho, Paulo C.; Gozzo, Fábio C.; Castleberry, Mark; Thompson, Thomas B.; Davidson, W. Sean

    2016-01-01

    Apolipoprotein (apo) A-I mediates many of the anti-atherogenic functions attributed to high density lipoprotein. Unfortunately, efforts toward a high resolution structure of full-length apoA-I have not been fruitful, although there have been successes with deletion mutants. Recently, a C-terminal truncation (apoA-IΔ185–243) was crystallized as a dimer. The structure showed two helical bundles connected by a long, curved pair of swapped helical domains. To compare this structure to that existing under solution conditions, we applied small angle x-ray scattering and isotope-assisted chemical cross-linking to apoA-IΔ185–243 in its dimeric and monomeric forms. For the dimer, we found evidence for the shared domains and aspects of the N-terminal bundles, but not the molecular curvature seen in the crystal. We also found that the N-terminal bundles equilibrate between open and closed states. Interestingly, this movement is one of the transitions proposed during lipid binding. The monomer was consistent with a model in which the long shared helix doubles back onto the helical bundle. Combined with the crystal structure, these data offer an important starting point to understand the molecular details of high density lipoprotein biogenesis. PMID:26755744

  16. Elevated homocysteine reduces apolipoprotein A-I expression in hyperhomocysteinemic mice and in males with coronary artery disease.

    PubMed

    Mikael, Leonie G; Genest, Jacques; Rozen, Rima

    2006-03-03

    Hyperhomocysteinemia, a risk factor for cardiovascular disease, is caused by nutritional or genetic disturbances in homocysteine metabolism. A polymorphism in methylenetetrahydrofolate reductase (MTHFR) is the most common genetic cause of mild hyperhomocysteinemia. To examine mechanisms by which an elevation in plasma homocysteine leads to vascular disease, we first performed microarray analyses in livers of Mthfr-deficient mice and identified differentially expressed genes that are involved in lipid and cholesterol metabolism. Microarrays and RT-PCR showed decreased mRNA for apolipoprotein A (ApoA)-IV and for ApoA-I and increased mRNA for cholesterol 7alpha hydroxylase (Cyp7A1) in Mthfr(+/-) mice compared with Mthfr(+/+) mice. Western blotting revealed that ApoA-I protein levels in liver and plasma of Mthfr(+/-) mice were 52% and 62% of levels in the respective tissues of Mthfr(+/+) mice. We also performed Western analysis for plasma ApoA-I protein levels in 60 males with coronary artery disease and identified a significant (P<0.01) negative correlation (-0.33) between ApoA-I and plasma homocysteine levels. This cohort also displayed a negative correlation (-0.24, P=0.06) between high-density lipoprotein cholesterol and plasma homocysteine. Treatment of HepG2 cells with supraphysiological levels of 5 mmol/L homocysteine reduced peroxisome proliferator-activated receptor (PPAR) alpha and ApoA-I protein levels and decreased ApoA-I promoter activity. Transfection with a PPARalpha construct upregulated ApoA-I and MTHFR. Our results suggest that hyperhomocysteinemia may increase risk of atherosclerosis by decreasing expression of ApoA-I and increasing expression of CYP7A1.

  17. Distinct Hepatic Receptors for Low Density Lipoprotein and Apolipoprotein E in Humans

    NASA Astrophysics Data System (ADS)

    Hoeg, Jeffrey M.; Demosky, Stephen J.; Gregg, Richard E.; Schaefer, Ernst J.; Brewer, H. Bryan

    1985-02-01

    Since the liver is a central organ for lipid and lipoprotein synthesis and catabolism, hepatic receptors for specific apolipoproteins on plasma lipoproteins would be expected to modulate lipid and lipoprotein metabolism. The role of hepatic receptors for low density lipoproteins and apolipoprotein E-containing lipoproteins was evaluated in patients with complementary disorders in lipoprotein metabolism: abetalipoproteinemia and homozygous familial hypercholesterolemia. In addition, hepatic membranes from a patient with familial hypercholesterolemia were studied and compared before and after portacaval shunt surgery. The results establish that the human liver has receptors for apolipoproteins B and E. Furthermore, in the human, hepatic receptors for low density lipoproteins and apolipoprotein E are genetically distinct and can undergo independent control.

  18. Effect of Body Mass Index on Apolipoprotein A-I Kinetics in Middle-Aged Men and Postmenopausal Women

    PubMed Central

    Welty, Francine K.; Lichtenstein, Alice H.; Lamon-Fava, Stefania; Schaefer, Ernst J.; Marsh, Julian B.

    2009-01-01

    The effect of body mass index (BMI) and obesity on apo A-I levels and kinetics was examined by gender. ApoA-I kinetics were determined with a primed-constant infusion of deuterated leucine in the fed state in 19 men and 13 postmenopausal women. Compared to nonobese men, nonobese women had a higher HDL-C and apoA-I level due to a 48% higher apoA-I PR (p=0.05). Obesity had no significant effects on apoA-I kinetics in women. In contrast, compared to non-obese men, obese men had a 9% lower apoA-I level due to a 64% higher FCR partially offset by a 47% higher PR. Obese women had a 52% higher HDL-C than obese men (50 mg/dL vs 33 mg/dL, respectively, p=0.012), a finding related to the faster apoA-I FCR in obese men. BMI was directly correlated with apoA-1 FCR (r = 0.84, p < 0.001) and PR (r = 0.79, p < 0.001) in men but not women. 62% of the variability in PR and 71% of the variability in FCR were due to BMI in men and only 3% and 23%, respectively, in women. In conclusion, BMI has a significant effect on apoA-I PR and FCR in men but not in women. PMID:17570251

  19. Levels and changes of HDL cholesterol and apolipoprotein A-I in relation to risk of cardiovascular events among statin-treated patients; a meta-analysis

    PubMed Central

    Boekholdt, S. Matthijs; Arsenault, Benoit J.; Hovingh, G. Kees; Mora, Samia; Pedersen, Terje R.; LaRosa, John C.; Welch, K.M.A.; Amarenco, Pierre; DeMicco, David A.; Tonkin, Andrew M.; Sullivan, David R.; Kirby, Adrienne; Colhoun, Helen M.; Hitman, Graham A.; Betteridge, D. John; Durrington, Paul N.; Clearfield, Michael B.; Downs, John R.; Gotto, Antonio M.; Ridker, Paul M.; Kastelein, John J.P.

    2013-01-01

    Background It is unclear whether levels of high-density lipoprotein cholesterol (HDL-C) or apolipoprotein A-I (apoA-I) remain inversely associated with cardiovascular risk among patients who achieve very low levels of low-density lipoprotein cholesterol (LDL-C) on statin therapy. It is also unknown whether a rise in HDL-C or apoA-I after initiation of statin therapy is associated with a reduced cardiovascular risk. Methods and results We performed a meta-analysis of 8 statin trials in which lipids and apolipoproteins were determined in all study participants at baseline and at 1-year follow-up. Individual patient data were obtained for 38,153 trial participants allocated to statin therapy, of whom 5387 suffered a major cardiovascular event. HDL-C levels were associated with a reduced risk of major cardiovascular events (adjusted hazard ratio 0.83, 95%CI 0.81–0.86 per 1 standard deviation increment), as were apoA-I levels (HR 0.79, 95%CI 0.72–0.82). This association was also observed among patients achieving on-statin LDL-C levels < 50 mg/dL. An increase of HDL-C was not associated with reduced cardiovascular risk (HR 0.98, 95%CI 0.94–1.01 per 1 standard deviation increment), whereas a rise in apoA-I was (HR 0.93, 95%CI 0.90–0.97). Conclusions Among patients treated with statin therapy, HDL-C and apoA-I levels were strongly associated with a reduced cardiovascular risk, even among those achieving very low LDL-C. An apoA-I increase was associated with a reduced risk of major cardiovascular events, whereas for HDL-C this was not the case. These findings suggest that therapies that increase apoA-I concentration require further exploration with regard to cardiovascular risk reduction. PMID:23965489

  20. In vivo PET imaging with [(18)F]FDG to explain improved glucose uptake in an apolipoprotein A-I treated mouse model of diabetes.

    PubMed

    Cochran, Blake J; Ryder, William J; Parmar, Arvind; Tang, Shudi; Reilhac, Anthonin; Arthur, Andrew; Charil, Arnaud; Hamze, Hasar; Barter, Philip J; Kritharides, Leonard; Meikle, Steven R; Gregoire, Marie-Claude; Rye, Kerry-Anne

    2016-09-01

    Type 2 diabetes is characterised by decreased HDL levels, as well as the level of apolipoprotein A-I (apoA-I), the main apolipoprotein of HDLs. Pharmacological elevation of HDL and apoA-I levels is associated with improved glycaemic control in patients with type 2 diabetes. This is partly due to improved glucose uptake in skeletal muscle. This study used kinetic modelling to investigate the impact of increasing plasma apoA-I levels on the metabolism of glucose in the db/db mouse model. Treatment of db/db mice with apoA-I for 2 h significantly improved both glucose tolerance (AUC 2574 ± 70 mmol/l × min vs 2927 ± 137 mmol/l × min, for apoA-I and PBS, respectively; p < 0.05) and insulin sensitivity (AUC 388.8 ± 23.8 mmol/l × min vs 194.1 ± 19.6 mmol/l × min, for apoA-I and PBS, respectively; p < 0.001). ApoA-I treatment also increased glucose uptake by skeletal muscle in both an insulin-dependent and insulin-independent manner as evidenced by increased uptake of fludeoxyglucose ([(18)F]FDG) from plasma into gastrocnemius muscle in apoA-I treated mice, both in the absence and presence of insulin. Kinetic modelling revealed an enhanced rate of insulin-mediated glucose phosphorylation (k 3) in apoA-I treated mice (3.5 ± 1.1 × 10(-2) min(-1) vs 2.3 ± 0.7 × 10(-2) min(-1), for apoA-I and PBS, respectively; p < 0.05) and an increased influx constant (3.7 ± 0.6 × 10(-3) ml min(-1) g(-1) vs 2.0 ± 0.3 × 10(-3) ml min(-1) g(-1), for apoA-I and PBS, respectively; p < 0.05). Treatment of L6 rat skeletal muscle cells with apoA-I for 2 h indicated that increased hexokinase activity mediated the increased rate of glucose phosphorylation. These findings indicate that apoA-I improves glucose disposal in db/db mice by improving insulin sensitivity and enhancing glucose phosphorylation.

  1. The Effect of Preoperative Apolipoprotein A-I on the Prognosis of Surgical Renal Cell Carcinoma: A Retrospective Large Sample Study.

    PubMed

    Guo, Shengjie; He, Xiaobo; Chen, Qian; Yang, Guangwei; Yao, Kai; Dong, Pei; Ye, Yunlin; Chen, Dong; Zhang, Zhiling; Qin, Zike; Liu, Zhuowei; Li, Zaishang; Xue, Yunfei; Zhang, Meng; Liu, Ruiwu; Zhou, Fangjian; Han, Hui

    2016-03-01

    The prognostic value of serum lipid-profile in renal cell cancer (RCC) remains unknown. The purpose of the study is to explore the association between the serum lipid-profile and RCC patients.The levels of preoperative serum lipid-profile (including cholesterol, triglycerides, high-density lipoprotein-cholesterol [HDL-C], low-density lipoprotein-cholesterol [LDL-C], apolipoprotein A-I [ApoA-I], and apolipoprotein B [ApoB]) were retrospectively performed in 786 patients with RCC. The cutoff values of the lipids were determined by the receiver-operating characteristic (ROC) curve analysis. Univariate and multivariate Cox regression analyses were performed to investigate the prognostic value of serum lipids in RCC.Combined ROC analysis and univariate and multivariate Cox regression analyses, for overall survival (OS), revealed patients with low ApoA-I (<1.04) had significantly lower OS than the high ApoA-I (≥1.04) group (multivariate Cox regression analyses, hazard ratio [HR], 0.57; P = 0.003). Not only in the whole RCC cohort but also in the subgroups stratified according to the pT1-2 (P = 0.002), pN0 (P < 0.001), and pM0 (P = 0.001) status, respectively. Moreover, in the 755 patients with nonmetastasis, the low ApoA-I group was also associated with shortened disease-free survival (DFS) time compared to the high ApoA-I group (multivariate Cox regression analyses, HR, 0.65; P = 0.033). However, the other lipids were not independent prognostic factors for surgical RCC.An elevated level of preoperative ApoA-I was demonstrated to be related with better survival in patients with surgical RCC. Measuring the preoperative ApoA-I might be a simple way for finding the poor prognostic patients who should enrolled in further clinical trials and management.

  2. Cholesteryl Ester Transfer Protein Inhibition With Anacetrapib Decreases Fractional Clearance Rates of High-Density Lipoprotein Apolipoprotein A-I and Plasma Cholesteryl Ester Transfer Protein.

    PubMed

    Reyes-Soffer, Gissette; Millar, John S; Ngai, Colleen; Jumes, Patricia; Coromilas, Ellie; Asztalos, Bela; Johnson-Levonas, Amy O; Wagner, John A; Donovan, Daniel S; Karmally, Wahida; Ramakrishnan, Rajasekhar; Holleran, Stephen; Thomas, Tiffany; Dunbar, Richard L; deGoma, Emil M; Rafeek, Hashmi; Baer, Amanda L; Liu, Yang; Lassman, Michael E; Gutstein, David E; Rader, Daniel J; Ginsberg, Henry N

    2016-05-01

    Anacetrapib (ANA), an inhibitor of cholesteryl ester transfer protein (CETP) activity, increases plasma concentrations of high-density lipoprotein cholesterol (HDL-C), apolipoprotein A-I (apoA)-I, apoA-II, and CETP. The mechanisms responsible for these treatment-related increases in apolipoproteins and plasma CETP are unknown. We performed a randomized, placebo (PBO)-controlled, double-blind, fixed-sequence study to examine the effects of ANA on the metabolism of HDL apoA-I and apoA-II and plasma CETP. Twenty-nine participants received atorvastatin (ATV) 20 mg/d plus PBO for 4 weeks, followed by ATV plus ANA 100 mg/d for 8 weeks (ATV-ANA). Ten participants received double PBO for 4 weeks followed by PBO plus ANA for 8 weeks (PBO-ANA). At the end of each treatment, we examined the kinetics of HDL apoA-I, HDL apoA-II, and plasma CETP after D3-leucine administration as well as 2D gel analysis of HDL subspecies. In the combined ATV-ANA and PBO-ANA groups, ANA treatment increased plasma HDL-C (63.0%; P<0.001) and apoA-I levels (29.5%; P<0.001). These increases were associated with reductions in HDL apoA-I fractional clearance rate (18.2%; P=0.002) without changes in production rate. Although the apoA-II levels increased by 12.6% (P<0.001), we could not discern significant changes in either apoA-II fractional clearance rate or production rate. CETP levels increased 102% (P<0.001) on ANA because of a significant reduction in the fractional clearance rate of CETP (57.6%, P<0.001) with no change in CETP production rate. ANA treatment increases HDL apoA-I and CETP levels by decreasing the fractional clearance rate of each protein. © 2016 American Heart Association, Inc.

  3. Modified apolipoprotein (apo) A-I by artificial sweetener causes severe premature cellular senescence and atherosclerosis with impairment of functional and structural properties of apoA-I in lipid-free and lipid-bound state.

    PubMed

    Jang, Wookju; Jeoung, Nam Ho; Cho, Kyung-Hyun

    2011-05-01

    Long-term consumption of artificial sweeteners (AS) has been the recent focus of safety concerns. However, the potential risk of the AS in cardiovascular disease and lipoprotein metabolism has not been investigated sufficiently. We compared the influence of AS (aspartame, acesulfame K, and saccharin) and fructose in terms of functional and structural correlations of apolipoprotein (apo) A-I and high-density lipoproteins (HDL), which have atheroprotective effects. Long-term treatment of apoA-I with the sweetener at physiological concentration (3 mM for 168 h) resulted in loss of antioxidant and phospholipid binding activities with modification of secondary structure. The AS treated apoA-I exhibited proteolytic cleavage to produce 26 kDa-fragment. They showed pro-atherogenic properties in acetylated LDL phagocytosis of macrophages. Each sweetener alone or sweetener-treated apoA-I caused accelerated senescence in human dermal fibroblasts. These results suggest that long-term consumption of AS might accelerate atherosclerosis and senescence via impairment of function and structure of apoA-I and HDL.

  4. Modified Apolipoprotein (apo) A-I by Artificial Sweetener Causes Severe Premature Cellular Senescence and Atherosclerosis with Impairment of Functional and Structural Properties of apoA-I in Lipid-Free and Lipid-Bound State

    PubMed Central

    Jang, Wookju; Jeoung, Nam Ho; Cho, Kyung-Hyun

    2011-01-01

    Long-term consumption of artificial sweeteners (AS) has been the recent focus of safety concerns. However, the potential risk of the AS in cardiovascular disease and lipoprotein metabolism has not been investigated sufficiently. We compared the influence of AS (aspartame, acesulfame K, and saccharin) and fructose in terms of functional and structural correlations of apolipoprotein (apo) A-I and high-density lipoproteins (HDL), which have atheroprotective effects. Long-term treatment of apoA-I with the sweetener at physiological concentration (3 mM for 168 h) resulted in loss of antioxidant and phospholipid binding activities with modification of secondary structure. The AS treated apoA-I exhibited proteolytic cleavage to produce 26 kDa-fragment. They showed pro-atherogenic properties in acetylated LDL phagocytosis of macrophages. Each sweetener alone or sweetener-treated apoA-I caused accelerated senescence in human dermal fibroblasts. These results suggest that long-term consumption of AS might accelerate atherosclerosis and senescence via impairment of function and structure of apoA-I and HDL. PMID:21533907

  5. An abundant dysfunctional apolipoprotein A1 in human atheroma

    PubMed Central

    Huang, Ying; DiDonato, Joseph A.; Levison, Bruce S.; Schmitt, Dave; Li, Lin; Wu, Yuping; Buffa, Jennifer; Kim, Timothy; Gerstenecker, Gary; Gu, Xiaodong; Kadiyala, Chandra; Wang, Zeneng; Culley, Miranda K.; Hazen, Jennie E.; DiDonato, Anthony J.; Fu, Xiaoming; Berisha, Stela; Peng, Daoquan; Nguyen, Truc; Liang, Shaohong; Chuang, Chia-Chi; Cho, Leslie; Plow, Edward F.; Fox, Paul L.; Gogonea, Valentin; Tang, W.H. Wilson; Parks, John S.; Fisher, Edward A.; Smith, Jonathan D.; Hazen, Stanley L.

    2014-01-01

    Recent studies indicate high density lipoproteins (HDL) and their major structural protein, apolipoprotein A1 (apoA1), recovered from human atheroma, are dysfunctional and extensively oxidized by myeloperoxidase (MPO), while in vitro oxidation of apoA1/HDL by MPO impairs its cholesterol acceptor function. We developed a high affinity monoclonal antibody (mAb) that specifically recognizes apoA1/HDL modified by the MPO/H2O2/Cl-system using phage display affinity maturation. An oxindolyl alanine (2-OH-Trp) moiety at tryptophan 72 of apoA1 is the immunogenic epitope. Mutagenesis studies confirm a critical role for apoA1 Trp72 in MPO-mediated inhibition of ABCA1-dependent cholesterol acceptor activity of apoA1 in vitro and in vivo. ApoA1 containing a 2-OH-Trp72 group (oxTrp72-apoA1) is in low abundance within the circulation, but accounts for 20% of the apoA1 in atherosclerotic plaque. OxTrp72-apoA1 recovered from human atheroma or plasma was lipid-poor, virtually devoid of cholesterol acceptor activity, and demonstrated both potent pro-inflammatory activities on endothelial cells and impaired HDL biogenesis activity in vivo. Elevated oxTrp72-apoA1 levels in subjects presenting to a cardiology clinic (n=627) were associated with increased cardiovascular disease risk. Circulating oxTrp72-apoA1 levels may serve as a way to monitor a pro-atherogenic process in the artery wall. PMID:24464187

  6. Specific Regional Transcription of Apolipoprotein E in Human Brain Neurons

    PubMed Central

    Xu, Pu-Ting; Gilbert, John R.; Qiu, Hui-Ling; Ervin, John; Rothrock-Christian, Tracie R.; Hulette, Christine; Schmechel, Donald E.

    1999-01-01

    In central nervous system injury and disease, apolipoprotein E (APOE, gene; apoE, protein) might be involved in neuronal injury and death indirectly through extracellular effects and/or more directly through intracellular effects on neuronal metabolism. Although intracellular effects could clearly be mediated by neuronal uptake of extracellular apoE, recent experiments in injury models in normal rodents and in mice transgenic for the human APOE gene suggest the additional possibility of intraneuronal synthesis. To examine whether APOE might be synthesized by human neurons, we performed in situ hybridization on paraffin-embedded and frozen brain sections from three nondemented controls and five Alzheimer’s disease (AD) patients using digoxigenin-labeled antisense and sense cRNA probes to human APOE. Using the antisense APOE probes, we found the expected strong hybridization signal in glial cells as well as a generally fainter signal in selected neurons in cerebral cortex and hippocampus. In hippocampus, many APOE mRNA-containing neurons were observed in sectors CA1 to CA4 and the granule cell layer of the dentate gyrus. In these regions, APOE mRNA containing neurons could be observed adjacent to nonhybridizing neurons of the same cell class. APOE mRNA transcription in neurons is regionally specific. In cerebellar cortex, APOE mRNA was seen only in Bergmann glial cells and scattered astrocytes but not in Purkinje cells or granule cell neurons. ApoE immunocytochemical localization in semi-adjacent sections supported the selectivity of APOE transcription. These results demonstrate the expected result that APOE mRNA is transcribed and expressed in glial cells in human brain. The important new finding is that APOE mRNA is also transcribed and expressed in many neurons in frontal cortex and human hippocampus but not in neurons of cerebellar cortex from the same brains. This regionally specific human APOE gene expression suggests that synthesis of apoE might play a role

  7. Apolipoprotein A-I Mimetic Peptide D-4F Reduces Cardiac Hypertrophy and Improves Apolipoprotein A-I-Mediated Reverse Cholesterol Transport From Cardiac Tissue in LDL Receptor-null Mice Fed a Western Diet.

    PubMed

    Han, Jie; Zhang, Song; Ye, Ping; Liu, Yong-Xue; Qin, Yan-Wen; Miao, Dong-Mei

    2016-05-01

    Epidemiological studies have suggested that hypercholesterolemia is an independent determinant of increased left ventricular (LV) mass. Because high-density lipoprotein and its major protein apolipoprotein A-I (apoA-I) mediate reverse cholesterol transport (RCT) and have cardiac protective effects, we hypothesized that the apoA-I mimetic peptide D-4F could promote RCT in cardiac tissue and decrease cardiac hypertrophy induced by hypercholesterolemia. Low-density lipoprotein receptor-null mice were fed by a Western diet for 18 weeks and then randomized to receive water, or D-4F 0.3 mg/mL, or D-4F 0.5 mg/mL added to drinking water for 6 weeks. After D-4F administration, an increase in high-density lipoprotein cholesterol and a decrease in low-density lipoprotein cholesterol, total cholesterol, and triglyceride in a trend toward dose-responsivity were found in cardiac tissue. Ultrasound biomicroscopy revealed a reduction in LV posterior wall end-diastolic dimension, and an increase in mitral valve E/A ratio and LV ejection fraction. Hematoxylin-eosin staining showed reduced LV wall thickness and myocardial cell diameter. The protein levels of ABCA1 and LXRα were elevated in cardiac tissue of D-4F treated mice compared with the controls (P < 0.05). These results demonstrated that D-4F treatment reduced cardiac hypertrophy, and improved cardiac performance in low-density lipoprotein receptor-null mice fed a Western diet, presumably through the LXRα-ABCA1 pathway associated with enhanced myocardial RCT.

  8. Heparin and Methionine Oxidation Promote the Formation of Apolipoprotein A-I Amyloid Comprising α-Helical and β-Sheet Structures.

    PubMed

    Townsend, David; Hughes, Eleri; Hussain, Rohanah; Siligardi, Giuliano; Baldock, Sarah; Madine, Jillian; Middleton, David A

    2017-03-21

    Peptides derived from apolipoprotein A-I (apoA-I), the main component of high-density lipoprotein (HDL), constitute the main component of amyloid deposits that colocalize with atherosclerotic plaques. Here we investigate the molecular details of full-length, lipid-deprived apoA-I after assembly into insoluble aggregates under physiologically relevant conditions known to induce aggregation in vitro. Unmodified apoA-I is shown to remain soluble at pH 7 for at least 3 days, retaining its native α-helical-rich structure. Upon acidification to pH 4, apoA-I rapidly assembles into insoluble nonfibrillar aggregates lacking the characteristic cross-β features of amyloid. In the presence of heparin, the rate and thioflavin T responsiveness of the aggregates formed at pH 4 increase and short amyloid-like fibrils are observed, which give rise to amyloid-characteristic X-ray reflections at 4.7 and 10 Å. Solid-state nuclear magnetic resonance (SSNMR) and synchrotron radiation circular dichroism spectroscopy of fibrils formed in the presence of heparin show they retain some α-helical characteristics together with new β-sheet structures. Interestingly, SSNMR indicates a similar molecular structure of aggregates formed in the absence of heparin at pH 6 after oxidation of the three methionine residues, although their morphology is rather different from that of the heparin-derived fibrils. We propose a model for apoA-I aggregation in which perturbations of a four-helix bundle-like structure, induced by interactions of heparin or methionine oxidation, cause the partially helical N-terminal residues to disengage from the remaining, intact helices, thereby allowing self-assembly via β-strand associations.

  9. Contribution of apolipoprotein A-I to the reduction in high-sensitivity C-reactive protein levels by different statins: comparative study of pitavastatin and atorvastatin.

    PubMed

    Tani, Shigemasa; Takahashi, Atsuhiko; Nagao, Ken; Hirayama, Atsushi

    2015-11-01

    Recently, investigation may have focused on modification of apolipoprotein A-I (apoA-I) associated with anti-inflammatory effect for the potential prevention of cardiovascular events. The purpose of this study was to evaluate the effects of atorvastatin and pitavastatin on serum apoA-I levels and to investigate the role of apoA-I in the anti-inflammatory effect of statin. We conducted a 6-month, prospective, randomized, open-label study in which we assigned hypercholesterolemic patients to a pitavastatin group (n = 52; 2 mg/day) or an atorvastatin group (n = 52; 10 mg/day) to investigate the effects of these two statins on the serum apoA-I levels and serum high-sensitivity C-reactive protein (hs-CRP) levels. There were no significant differences between the two groups in the changes in the low-density lipoprotein cholesterol, high-density lipoprotein cholesterol (HDL-C), or hs-CRP levels, but the change in apoA-I in the pitavastatin group was significantly greater than in the atorvastatin group (5.3 vs. 1.4 %; p = 0.0001). A stepwise regression analysis revealed that the percent change in (Δ) serum apoA-I level was an independent predictor of the Δ serum hs-CRP (standard correlation coefficient = -0.198; p = 0.047). However, there was a significant negative correlation between the Δ apoA-I levels and Δ hs-CRP levels in the pitavastatin group (r = -0.283, p = 0.042), but not the atorvastatin group (r = -0.133, p = 0.356). The results suggest that the contribution of apoA-I to the reduction in serum hs-CRP levels by these two statins may be different. A decrease in hs-CRP level accompanied by an increase in apoA-I level may be involved in the pleiotropic effects of pitavastatin.

  10. High level of serum apolipoprotein A-I is a favorable prognostic factor for overall survival in esophageal squamous cell carcinoma.

    PubMed

    Wang, Xue-Ping; Li, Xiao-Hui; Zhang, Lin; Lin, Jian-Hua; Huang, Hao; Kang, Ting; Mao, Min-Jie; Chen, Hao; Zheng, Xin

    2016-07-21

    Noninvasive prognostic tools for esophageal squamous cell carcinoma (ESCC) are urgently needed. Serum lipids and lipoproteins are used for the prognosis of certain diseases; however, the prognostic value of serum apolipoprotein A-I (ApoA-I) in ESCC has not been described. Pre-treatment serum lipids and lipoprotein concentrations (including ApoA-I, Apo-B, HDL-C, LDL-C, TC and TG) were analyzed retrospectively and compared between 210 patients with ESCC and 219 healthy controls. The prognostic significance of serum lipids and lipoproteins was determined by univariate and multivariate Cox hazard models in ESCC. Clinical characteristics (age, sex, pT status, pN status, pM status, pTNM status, histological differentiation or alcohol index) had no influence on baseline ApoA-I level. Serum ApoA-I, HDL-C, LDL-C, and TC levels were significantly lower and Apo-B was significantly higher in ESCC patients than in normal controls. On univariate analysis, ApoA-I, alcohol index, pT status, pN status and pTNM status were associated with significantly poor survival, and ApoA-I (p = 0.039), alcohol index (p = 0.037) and pTNM status (p = 0.000) were identified as prognostic factors associated with shorter survival in the multivariate analysis. Overall survival was shorter in ESCC patients with decreased pre-treatment ApoA-I levels. Our findings suggest that serum ApoA-I level should be evaluated as a predictor of survival in patients with ESCC.

  11. ITRAQ-based quantitative proteomics reveals apolipoprotein A-I and transferrin as potential serum markers in CA19-9 negative pancreatic ductal adenocarcinoma.

    PubMed

    Lin, Chao; Wu, Wen-Chuan; Zhao, Guo-Chao; Wang, Dan-Song; Lou, Wen-Hui; Jin, Da-Yong

    2016-08-01

    Currently the diagnosis of pancreatic ductal adenocarcinoma (PDAC) relies on CA19-9 and radiological means, whereas some patients do not have elevated levels of CA19-9 secondary to pancreatic cancer. The purpose of this study was to identify potential serum biomarkers for CA19-9 negative PDAC.A total of 114 serum samples were collected from 3 groups: CA19-9 negative PDAC patients (n = 34), CA19-9 positive PDAC patients (n = 44), and healthy volunteers (n = 36), whereas the first 12 samples from each group were used for isobaric tags for relative and absolute quantitation (iTRAQ) analysis. Thereafter, candidate biomarkers were selected for validation by enzyme-linked immunosorbent assay (ELISA) with the rest specimens.Using the iTRAQ approach, a total of 5 proteins were identified as significantly different between CA19-9 negative PDAC patients and healthy subjects according to our defined criteria. Apolipoprotein A-I (APOA-I) and transferrin (TF) were selected to validate the proteomic results by ELISA in a further 78 serum specimens. It revealed that TF significantly correlated with the degree of histological differentiation (P = 0.042), and univariate and multivariate analyses indicated that TF is an independent prognostic factor for survival (hazard ratio, 0.302; 95% confidence interval, 0.118-0.774; P = 0.013) of patients with PDAC after curative surgery.ITRAQ-based quantitative proteomics revealed that APOA-I and TF may be potential CA19-9 negative PDAC serum markers.

  12. Stimulation of Hepatic Apolipoprotein A-I Production by Novel Thieno-Triazolodiazepines: Roles of the Classical Benzodiazepine Receptor, PAF Receptor, and Bromodomain Binding

    PubMed Central

    Kempen, Herman J; Bellus, Daniel; Fedorov, Oleg; Nicklisch, Silke; Filippakopoulos, Panagis; Picaud, Sarah; Knapp, Stefan

    2013-01-01

    Expression and secretion of apolipoprotein A-I (apoA-I) by cultured liver cells can be markedly stimulated by triazolodiazepines (TZDs). It has been shown previously that the thieno-TZD Ro 11-1464 increases plasma levels of apoA-I and in vivomacrophage reverse cholesterol transport in mice. However, these effects were only seen at high doses, at which the compound could act on central benzodiazepine (BZD) receptors or platelet activating factor (PAF) receptors, interfering with its potential utility. In this work, we describe 2 new thieno-TZDs MDCO-3770 and MDCO-3783, both derived from Ro 11-1464. These compounds display the same high efficacy on apoA-I production, metabolic stability, and lack of cytotoxicity in cultured hepatocytes as Ro 11-1464, but they do not bind to the central BZD receptor and PAF receptor. The quinazoline RVX-208 was less efficacious in stimulating apoA-I production and displayed signs of cytotoxicity. Certain TZDs stimulating apoA-I production are now known to be inhibitors of bromodomain (BRD) extra-terminal (BET) proteins BRDT, BRD2, BRD3, and BRD4, and this inhibition was inferred as a main molecular mechanism for their effect on apoA-I expression. We show here that the thieno-TZD (+)-JQ1, a potent BET inhibitor, strongly stimulated apoA-I production in Hep-G2 cells, but that its enantiomer (−)-JQ1, which has no BET inhibitor activity, also showed considerable effect on apoA-I production. MDCO-3770 and MDCO-3783 also inhibited BRD3 and BRD4 in vitro, with potency somewhat below that of (+)-JQ1. We conclude that the effect of thieno-TZDs on apoA-I expression is not due to inhibition of the BZD or PAF receptors and is not completely explained by transcriptional repression by BET proteins. PMID:25278768

  13. Stimulation of Hepatic Apolipoprotein A-I Production by Novel Thieno-Triazolodiazepines: Roles of the Classical Benzodiazepine Receptor, PAF Receptor, and Bromodomain Binding.

    PubMed

    Kempen, Herman J; Bellus, Daniel; Fedorov, Oleg; Nicklisch, Silke; Filippakopoulos, Panagis; Picaud, Sarah; Knapp, Stefan

    2013-01-01

    Expression and secretion of apolipoprotein A-I (apoA-I) by cultured liver cells can be markedly stimulated by triazolodiazepines (TZDs). It has been shown previously that the thieno-TZD Ro 11-1464 increases plasma levels of apoA-I and in vivomacrophage reverse cholesterol transport in mice. However, these effects were only seen at high doses, at which the compound could act on central benzodiazepine (BZD) receptors or platelet activating factor (PAF) receptors, interfering with its potential utility. In this work, we describe 2 new thieno-TZDs MDCO-3770 and MDCO-3783, both derived from Ro 11-1464. These compounds display the same high efficacy on apoA-I production, metabolic stability, and lack of cytotoxicity in cultured hepatocytes as Ro 11-1464, but they do not bind to the central BZD receptor and PAF receptor. The quinazoline RVX-208 was less efficacious in stimulating apoA-I production and displayed signs of cytotoxicity. Certain TZDs stimulating apoA-I production are now known to be inhibitors of bromodomain (BRD) extra-terminal (BET) proteins BRDT, BRD2, BRD3, and BRD4, and this inhibition was inferred as a main molecular mechanism for their effect on apoA-I expression. We show here that the thieno-TZD (+)-JQ1, a potent BET inhibitor, strongly stimulated apoA-I production in Hep-G2 cells, but that its enantiomer (-)-JQ1, which has no BET inhibitor activity, also showed considerable effect on apoA-I production. MDCO-3770 and MDCO-3783 also inhibited BRD3 and BRD4 in vitro, with potency somewhat below that of (+)-JQ1. We conclude that the effect of thieno-TZDs on apoA-I expression is not due to inhibition of the BZD or PAF receptors and is not completely explained by transcriptional repression by BET proteins.

  14. ITRAQ-based quantitative proteomics reveals apolipoprotein A-I and transferrin as potential serum markers in CA19-9 negative pancreatic ductal adenocarcinoma

    PubMed Central

    Lin, Chao; Wu, Wen-Chuan; Zhao, Guo-Chao; Wang, Dan-Song; Lou, Wen-Hui; Jin, Da-Yong

    2016-01-01

    Abstract Currently the diagnosis of pancreatic ductal adenocarcinoma (PDAC) relies on CA19-9 and radiological means, whereas some patients do not have elevated levels of CA19-9 secondary to pancreatic cancer. The purpose of this study was to identify potential serum biomarkers for CA19-9 negative PDAC. A total of 114 serum samples were collected from 3 groups: CA19-9 negative PDAC patients (n = 34), CA19-9 positive PDAC patients (n = 44), and healthy volunteers (n = 36), whereas the first 12 samples from each group were used for isobaric tags for relative and absolute quantitation (iTRAQ) analysis. Thereafter, candidate biomarkers were selected for validation by enzyme-linked immunosorbent assay (ELISA) with the rest specimens. Using the iTRAQ approach, a total of 5 proteins were identified as significantly different between CA19-9 negative PDAC patients and healthy subjects according to our defined criteria. Apolipoprotein A-I (APOA-I) and transferrin (TF) were selected to validate the proteomic results by ELISA in a further 78 serum specimens. It revealed that TF significantly correlated with the degree of histological differentiation (P = 0.042), and univariate and multivariate analyses indicated that TF is an independent prognostic factor for survival (hazard ratio, 0.302; 95% confidence interval, 0.118–0.774; P = 0.013) of patients with PDAC after curative surgery. ITRAQ-based quantitative proteomics revealed that APOA-I and TF may be potential CA19-9 negative PDAC serum markers. PMID:27495108

  15. Effect of niacin on high-density lipoprotein apolipoprotein A-I kinetics in statin-treated patients with type 2 diabetes mellitus.

    PubMed

    Pang, Jing; Chan, Dick C; Hamilton, Sandra J; Tenneti, Vijay S; Watts, Gerald F; Barrett, P Hugh R

    2014-02-01

    To investigate the effect of extended-release (ER) niacin on the metabolism of high-density lipoprotein (HDL) apolipoprotein A-I (apoA-I) in men with type 2 diabetes mellitus on a background of optimal statin therapy. Twelve men with type 2 diabetes mellitus were recruited for a randomized, crossover design trial. Patients were randomized to rosuvastatin or rosuvastatin plus ER niacin for 12 weeks and then crossed over to the alternate therapy after a 3-week washout period. Metabolic studies were performed at the end of each treatment period. HDL apoA-I kinetics were measured after a standardized liquid mixed meal and a bolus injection of d3-leucine for 96 hours. Compartmental analysis was used to model the data. ER niacin significantly decreased plasma triglyceride, plasma cholesterol, non-HDL cholesterol, low-density lipoprotein cholesterol, and apoB (all P<0.05) and significantly increased HDL cholesterol and apoA-I concentrations (P<0.005 and P<0.05, respectively). ER niacin also significantly increased HDL apoA-I pool size (6,088 ± 292 versus 5,675 ± 305 mg; P<0.001), and this was attributed to a lower HDL apoA-I fractional catabolic rate (0.33 ± 0.01 versus 0.37 ± 0.02 pools/d; P<0.005), with no significant changes in HDL apoA-I production (20.93 ± 0.63 versus 21.72 ± 0.85 mg/kg per day; P=0.28). ER niacin increases HDL apoA-I concentration in statin-treated subjects with type 2 diabetes mellitus by lowering apoA-I fractional catabolic rate. The effect on HDL metabolism was independent of the reduction in plasma triglyceride with ER niacin treatment. Whether this finding applies to other dyslipidemic populations remains to be investigated.

  16. Effects of the Iowa and Milano Mutations on Apolipoprotein A-I Structure and Dynamics Determined by Hydrogen Exchange and Mass Spectrometry

    PubMed Central

    Chetty, Palaniappan Sevugan; Ohshiro, Maki; Saito, Hiroyuki; Dhanasekaran, Padmaja; Lund-Katz, Sissel; Mayne, Leland; Englander, Walter; Phillips, Michael C.

    2012-01-01

    The Iowa point mutation in apolipoprotein A-I (G26R; apoA-IIowa) leads to a systemic amyloidosis condition and the Milano mutation (R173C; apoA-IMil) is associated with hypoalphalipoproteinemia, a reduced plasma level of high density lipoprotein. To probe the structural effects that lead to these outcomes, we used amide hydrogen-deuterium exchange coupled with a fragment separation/mass spectrometry analysis (HX MS). The Iowa mutation inserts an arginine residue into the non-polar face of an α-helix that spans residues 7–44 and causes changes in structure and structural dynamics. This helix unfolds and other helices in the N-terminal helix bundle domain are destabilized. The segment encompassing residues 116–158, largely unstructured in wild type apoA-I, becomes helical. The helix spanning residues 81 to 115 is destabilized by 2 kcal/mol, increasing the small fraction of time it is transiently unfolded to 1% or more, which allows proteolysis at residue 83 in vivo over time, releasing an amyloid-forming peptide. The Milano mutation situated on the polar face of the helix spanning residues 147–178 destabilizes the helix bundle domain only moderately, but enough to allow cysteine-mediated dimerization which leads to the altered functionality of this variant. These results show how the HX MS approach can provide a powerful means for monitoring, in a non-perturbing way and at close to amino acid resolution, the structural, dynamic, and energetic consequences of biologically interesting point mutations. PMID:23066790

  17. β3-Adrenoceptor activation upregulates apolipoprotein A-I expression in HepG2 cells, which might further promote cholesterol efflux from macrophage foam cells

    PubMed Central

    Gao, Xia-qing; Li, Yan-fang; Jiang, Zhi-li

    2017-01-01

    Objective The aim of this study was to explore the effects of β3-adrenoceptor (β3-AR) activation on HepG2 cells and its influence on cholesterol efflux from macrophage foam cells. Materials and methods HepG2 cells were cultured and treated with the β3-AR agonist, BRL37344, and antagonist, SR52390A, and the expression of apolipoprotein (Apo) A-I, ApoA-II, ApoB, and β3-AR in the supernatants and cells was determined. The expression of peroxisome proliferator-activated receptor (PPAR) γ and PPARα in the HepG2 cells was also assessed. Next, using the RAW264.7 macrophage foam cell model, we also assessed the influence of the HepG2 cell supernatants on lipid efflux. The cholesterol content of the foam cells was also measured, and the cholesterol efflux from the macrophages was examined by determining 3H-labeled cholesterol levels. Expression of ATP-binding cassette transporter (ABC) A1 and ABCG1 of the macrophage foam cells was also assessed. Results β3-AR activation increased ApoA-I expression in both the HepG2 cells and the supernatants; PPARγ expression was upregulated, but PPARα expression was not. Treatment with GW9662 abolished the increased expression of ApoA-I induced by the β3-AR agonist. The HepG2 cell supernatants decreased the lipid accumulation and increased the cholesterol efflux from the macrophage foam cells. ABCA1 expression, but not ABCG1 expression, increased in the macrophage foam cells treated with BRL37344-treated HepG2 cell supernatants. Conclusion Activation of β3-AR in HepG2 cells upregulates ApoA-I expression, which might further promote cholesterol efflux from macrophage foam cells. PPARγ might be required for the induction of ApoA-I expression. PMID:28424539

  18. β3-Adrenoceptor activation upregulates apolipoprotein A-I expression in HepG2 cells, which might further promote cholesterol efflux from macrophage foam cells.

    PubMed

    Gao, Xia-Qing; Li, Yan-Fang; Jiang, Zhi-Li

    2017-01-01

    The aim of this study was to explore the effects of β3-adrenoceptor (β3-AR) activation on HepG2 cells and its influence on cholesterol efflux from macrophage foam cells. HepG2 cells were cultured and treated with the β3-AR agonist, BRL37344, and antagonist, SR52390A, and the expression of apolipoprotein (Apo) A-I, ApoA-II, ApoB, and β3-AR in the supernatants and cells was determined. The expression of peroxisome proliferator-activated receptor (PPAR) γ and PPARα in the HepG2 cells was also assessed. Next, using the RAW264.7 macrophage foam cell model, we also assessed the influence of the HepG2 cell supernatants on lipid efflux. The cholesterol content of the foam cells was also measured, and the cholesterol efflux from the macrophages was examined by determining (3)H-labeled cholesterol levels. Expression of ATP-binding cassette transporter (ABC) A1 and ABCG1 of the macrophage foam cells was also assessed. β3-AR activation increased ApoA-I expression in both the HepG2 cells and the supernatants; PPARγ expression was upregulated, but PPARα expression was not. Treatment with GW9662 abolished the increased expression of ApoA-I induced by the β3-AR agonist. The HepG2 cell supernatants decreased the lipid accumulation and increased the cholesterol efflux from the macrophage foam cells. ABCA1 expression, but not ABCG1 expression, increased in the macrophage foam cells treated with BRL37344-treated HepG2 cell supernatants. Activation of β3-AR in HepG2 cells upregulates ApoA-I expression, which might further promote cholesterol efflux from macrophage foam cells. PPARγ might be required for the induction of ApoA-I expression.

  19. Kinetic and thermodynamic analyses of spontaneous exchange between high-density lipoprotein-bound and lipid-free apolipoprotein A-I.

    PubMed

    Handa, Daisuke; Kimura, Hitoshi; Oka, Tatsuya; Takechi, Yuki; Okuhira, Keiichiro; Phillips, Michael C; Saito, Hiroyuki

    2015-02-03

    It is thought that apolipoprotein A-I (apoA-I) spontaneously exchanges between high-density lipoprotein (HDL)-bound and lipid-free states, which is relevant to the occurrence of preβ-HDL particles in plasma. To improve our understanding of the mechanistic basis for this phenomenon, we performed kinetic and thermodynamic analyses for apoA-I exchange between discoidal HDL-bound and lipid-free forms using fluorescence-labeled apoA-I variants. Gel filtration experiments demonstrated that addition of excess lipid-free apoA-I to discoidal HDL particles promotes exchange of apoA-I between HDL-associated and lipid-free pools without alteration of the steady-state HDL particle size. Kinetic analysis of time-dependent changes in NBD fluorescence upon the transition of NBD-labeled apoA-I from HDL-bound to lipid-free state indicates that the exchange kinetics are independent of the collision frequency between HDL-bound and lipid-free apoA-I, in which the lipid binding ability of apoA-I affects the rate of association of lipid-free apoA-I with the HDL particles and not the rate of dissociation of HDL-bound apoA-I. Thus, C-terminal truncations or mutations that reduce the lipid binding affinity of apoA-I strongly impair the transition of lipid-free apoA-I to the HDL-bound state. Thermodynamic analysis of the exchange kinetics demonstrated that the apoA-I exchange process is enthalpically unfavorable but entropically favorable. These results explain the thermodynamic basis of the spontaneous exchange reaction of apoA-I associated with HDL particles. The altered exchangeability of dysfunctional apoA-I would affect HDL particle rearrangement, leading to perturbed HDL metabolism.

  20. Nucleotide sequence and structure of the human apolipoprotein E gene.

    PubMed Central

    Paik, Y K; Chang, D J; Reardon, C A; Davies, G E; Mahley, R W; Taylor, J M

    1985-01-01

    The gene for human apolipoprotein E (apo-E) was selected from a library of cloned genomic DNA by screening with a specific cDNA hybridization probe, and its structure was characterized. The complete nucleotide sequence of the gene as well as 856 nucleotides of the 5' flanking region and 629 nucleotides of the 3' flanking region were determined. Analysis of the sequence showed that the mRNA-encoding region of the apo-E gene consists of four exons separated by three introns. In comparison to the structure of the mRNA, the introns are located in the 5' noncoding region, in the codon for glycine at position -4 of the signal peptide region, and in the codon for arginine at position +61 of the mature protein. The overall lengths of the apo-E gene and its corresponding mRNA are 3597 and 1163 nucleotides, respectively; a mature plasma protein of 299 amino acids is produced by this gene. Examination of the 5' terminus of the gene by S1 nuclease mapping shows apparent multiple transcription initiation sites. The proximal 5' flanking region contains a "TATA box" element as well as two nearby inverted repeat elements. In addition, there are four Alu family sequences associated with the apo-E gene: an Alu sequence located near each end of the gene and two Alu sequences located in the second intron. This knowledge of the structure permits a molecular approach to characterizing the regulation of the apo-E gene. Images PMID:2987927

  1. Apolipoprotein AI tertiary structures determine stability and phospholipid-binding activity of discoidal high-density lipoprotein particles of different sizes

    SciTech Connect

    Chen, Bin; Ren, Xuefeng; Neville, Tracey; Jerome, W. Gray; Hoyt, David W.; Sparks, Daniel L.; Ren, Gang; Wang, Jianjun

    2009-05-18

    Human high-density lipoprotein (HDL) plays a key role in the reverse cholesterol transport pathway that delivers excess cholesterol back to the liver for clearance. In vivo, HDL particles vary in size, shape and biological function. The discoidal HDL is a 140-240 kDa, disk-shaped intermediate of mature HDL. During mature spherical HDL formation, discoidal HDLs play a key role in loading cholesterol ester onto the HDL particles by activating the enzyme, lecithin:cholesterol acyltransferase (LCAT). One of the major problems for high-resolution structural studies of discoidal HDL is the difficulty in obtaining pure and, foremost, homogenous sample. We demonstrate here that the commonly used cholate dialysis method for discoidal HDL preparation usually contains 5-10% lipid-poor apoAI that significantly interferes with the high-resolution structural analysis of discoidal HDL using biophysical methods. Using an ultracentrifugation method, we quickly removed lipid-poor apoAI. We also purified discoidal reconstituted HDL (rHDL) into two pure discoidal HDL species of different sizes that are amendable for high-resolution structural studies. A small rHDL has a diameter of 7.6 nm, and a large rHDL has a diameter of 9.8 nm. We show that these two different sizes of discoidal HDL particles display different stability and phospholipid-binding activity. Interestingly, these property/functional differences are independent from the apoAI -helical secondary structure, but are determined by the tertiary structural difference of apoAI on different discoidal rHDL particles, as evidenced by two-dimensional NMR and negative stain electron microscopy data. Our result further provides the first high-resolution NMR data, demonstrating a promise of structural determination of discoidal HDL at atomic resolution using a combination of NMR and other biophysical techniques.

  2. Therapeutic ultrasound: Increased HDL-Cholesterol following infusions of acoustic microspheres and apolipoprotein A-I plasmids.

    PubMed

    Castle, Jason W; Kent, Kevin P; Fan, Ying; Wallace, Kirk D; Davis, Cynthia E L; Roberts, Jeannette C; Marino, Michael E; Thomenius, Kai E; Lim, Hae W; Coles, Eric; Davidson, Michael H; Feinstein, Steven B; DeMaria, Anthony

    2015-07-01

    Low levels of HDL-C are an independent cardiovascular risk factor associated with increased premature cardiovascular death. However, HDL-C therapies historically have been limited by issues relating to immunogenicity, hepatotoxicity and scalability, and have been ineffective in clinical trials. We examined the feasibility of using injectable acoustic microspheres to locally deliver human ApoA-I DNA plasmids in a pre-clinical model and quantify increased production of HDL-C in vivo. Our novel site-specific gene delivery system was examined in naïve rat model and comprised the following steps: (1) intravenous co-administration of a solution containing acoustically active microspheres (Optison™, GE Healthcare, Princeton, New Jersey) and human ApoA-I plasmids; (2) ultrasound verification of the presence of the microspheres within the liver vasculature; (3) External application of locally-directed acoustic energy, (4) induction of microsphere disruption and in situ sonoporation; (4) ApoA-I plasmid hepatic uptake; (5) transcription and expression of human ApoA-I protein; and (6) elevation of serum HDL-C. Co-administration of ApoA-I plasmids and acoustic microspheres, activated by external ultrasound energy, resulted in transcription and production of human ApoA-I protein and elevated serum HDL-C in rats (up to 61%; p-value < 0.05). HDL-C was increased in rats following ultrasound directed delivery of human ApoA-I plasmids by microsphere sonoporation. The present method provides a novel approach to promote ApoA-I synthesis and nascent HDL-C elevation, potentially permitting the use of a minimally-invasive ultrasound-based, gene delivery system for treating individuals with low HDL-C. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  3. Influence of C-terminal α-helix hydrophobicity and aromatic amino acid content on apolipoprotein A-I functionality.

    PubMed

    Lyssenko, Nicholas N; Hata, Mami; Dhanasekaran, Padmaja; Nickel, Margaret; Nguyen, David; Chetty, Palaniappan Sevugan; Saito, Hiroyuki; Lund-Katz, Sissel; Phillips, Michael C

    2012-03-01

    The apoA-I molecule adopts a two-domain tertiary structure and the properties of these domains modulate the ability to form HDL particles. Thus, human apoA-I differs from mouse apoA-I in that it can form smaller HDL particles; the C-terminal α-helix is important in this process and human apoA-I is unusual in containing aromatic amino acids in the non-polar face of this amphipathic α-helix. To understand the influence of these aromatic amino acids and the associated high hydrophobicity, apoA-I variants were engineered in which aliphatic amino acids were substituted with or without causing a decrease in overall hydrophobicity. The variants human apoA-I (F225L/F229A/Y236A) and apoA-I (F225L/F229L/A232L/Y236L) were compared to wild-type (WT) apoA-I for their abilities to (1) solubilize phospholipid vesicles and form HDL particles of different sizes, and (2) mediate cellular cholesterol efflux and create nascent HDL particles via ABCA1. The loss of aromatic residues and concomitant decrease in hydrophobicity in apoA-I (F225L/F229A/Y236A) has no effect on protein stability, but reduces by a factor of about three the catalytic efficiencies (V(max)/K(m)) of vesicle solubilization and cholesterol efflux; also, relatively large HDL particles are formed. With apoA-I (F225L/F229L/A232L/Y236L) where the hydrophobicity is restored by the presence of only leucine residues in the helix non-polar face, the catalytic efficiencies of vesicle solubilization and cholesterol efflux are similar to those of WT apoA-I; this variant forms smaller HDL particles. Overall, the results show that the hydrophobicity of the non-polar face of the C-terminal amphipathic α-helix plays a critical role in determining apoA-I functionality but aromatic amino acids are not required. This article is part of a Special Issue entitled Advances in High Density Lipoprotein Formation and Metabolism: A Tribute to John F. Oram (1945-2010). Copyright © 2011 Elsevier B.V. All rights reserved.

  4. PEGylated helper-dependent adenoviral vector expressing human Apo A-I for gene therapy in LDLR-deficient mice.

    PubMed

    Leggiero, E; Astone, D; Cerullo, V; Lombardo, B; Mazzaccara, C; Labruna, G; Sacchetti, L; Salvatore, F; Croyle, M; Pastore, L

    2013-12-01

    Helper-dependent adenoviral (HD-Ad) vectors have great potential for gene therapy applications; however, their administration induces acute toxicity that impairs safe clinical applications. We previously observed that PEGylation of HD-Ad vectors strongly reduces the acute response in murine and primate models. To evaluate whether PEGylated HD-Ad vectors combine reduced toxicity with the correction of pathological phenotypes, we administered an HD-Ad vector expressing the human apolipoprotein A-I (hApoA-I) to low-density lipoprotein (LDL)-receptor-deficient mice (a model for familial hypercholesterolemia) fed a high-cholesterol diet. Mice were treated with high doses of HD-Ad-expressing apo A-I or its PEGylated version. Twelve weeks later, LDL levels were lower and high-density lipoprotein (HDL) levels higher in mice treated with either of the vectors than in untreated mice. After terminal killing, the areas of atherosclerotic plaques were much smaller in the vector-treated mice than in the control animals. Moreover, the increase in pro-inflammatory cytokines was lower and consequently the toxicity profile better in mice treated with PEGylated vector than in mice treated with the unmodified vector. This finding indicates that the reduction in toxicity resulting from PEGylation of HD-Ad vectors does not impair the correction of pathological phenotypes. It also supports the clinical potential of these vectors for the correction of genetic diseases.

  5. The secondary structure of apolipoproteins in human HDL3 particles after chemical modification of their tyrosine, lysine, cysteine or arginine residues. A Fourier transform infrared spectroscopy study.

    PubMed

    Herzyk, E; Owen, J S; Chapman, D

    1988-09-02

    Fourier transform infrared spectra of apolipoprotein E-depleted human HDL3 have been obtained in H2O and 2H2O buffers. The absorption bands in the protein amide I and amide II regions (1700-1500 cm-1) were assigned to alpha-helical, disordered and beta-strand/beta-turn structures of apolipoproteins A-I and A-II (apoA-I and apoA-II), the apolipoprotein constituents of HDL3. Modification of HDL3 by tetranitromethane (TNM) treatment, acetylation, reduction plus alkylation and 1,2-cyclohexanedione treatment derivatised tyrosine, lysine, cysteine and arginine residues, respectively, and caused alteration of the secondary structure of the HDL3 apolipoproteins to different extents. Each of the chemical modifications caused changes in the frequency of bands associated with beta-strands/beta-turns, but only TNM treatment of HDL3, as judged by the second- and fourth-derivative spectra, resulted in a shift of the band assigned to the alpha-helical structure of the proteins. In agreement with other workers, only TNM treatment of HDL3 particles was found to inhibit their binding by high-affinity cell membrane receptors. It is proposed, therefore, that receptor recognition of HDL3 particles is dependent on conservation of the alpha-helix structures within apoA-I and apoA-II, and that beta-strand/beta-turn structures are not involved. This conclusion is consistent with the predominance of amphipathic alpha-helical structures in both apolipoproteins and with the relaxed specificity of the receptors which are thought to recognise both apoA-I and apoA-II.

  6. A novel truncated form of apolipoprotein A-I transported by dense LDL is increased in diabetic patients1[S

    PubMed Central

    Cubedo, Judit; Padró, Teresa; García-Arguinzonis, Maisa; Vilahur, Gemma; Miñambres, Inka; Pou, Jose María; Ybarra, Juan; Badimon, Lina

    2015-01-01

    Diabetic (DM) patients have exacerbated atherosclerosis and high CVD burden. Changes in lipid metabolism, lipoprotein structure, and dysfunctional HDL are characteristics of diabetes. Our aim was to investigate whether serum ApoA-I, the main protein in HDL, was biochemically modified in DM patients. By using proteomic technologies, we have identified a 26 kDa ApoA-I form in serum. MS analysis revealed this 26 kDa form as a novel truncated variant lacking amino acids 1-38, ApoA-IΔ(1-38). DM patients show a 2-fold increase in ApoA-IΔ(1-38) over nondiabetic individuals. ApoA-IΔ(1-38) is found in LDL, but not in VLDL or HDL, with an increase in LDL3 and LDL4 subfractions. To identify candidate mechanisms of ApoA-I truncation, we investigated potentially involved enzymes by in silico data mining, and tested the most probable molecule in an established animal model of diabetes. We have found increased hepatic cathepsin D activity as one of the potential proteases involved in ApoA-I truncation. Cathepsin D-cleaved ApoA-I exhibited increased LDL binding affinity and decreased antioxidant activity against LDL oxidation. In conclusion, we show for the first time: a) presence of a novel truncated ApoA-I form, ApoA-IΔ(1-38), in human serum; b) ApoA-IΔ(1-38) is transported by LDL; c) ApoA-IΔ(1-38) is increased in dense LDL fractions of DM patients; and d) cathepsin D-ApoA-I truncation may lead to ApoA-IΔ(1-38) binding to LDLs, increasing their susceptibility to oxidation and contributing to the high cardiovascular risk of DM patients. PMID:26168996

  7. Kinetic analysis of apolipoproteins in postprandial hypertriglyceridaemia rabbits.

    PubMed

    Hata, M; Ito, T; Ohwada, K

    2009-04-01

    The postprandial hypertriglyceridaemia (PHT) rabbit, developed as a new animal model of metabolic syndrome, is characterized by PHT, central obesity and glucose intolerance. For detailed investigation of lipid metabolism characteristics in PHT rabbit, the plasma levels of apolipoproteins A-I, B, C-II, C-III and E were measured. Movements of apolipoproteins B100 and B48 were investigated using sodium dodecyl sulphate-polyacrylamide gel electrophoresis to determine whether postprandially increased triglyceride is exogenous or endogenous. The level of apolipoproteins A-I, B, C-II and E were increased in PHT rabbit after feeding. Apolipoproteins B100 and B48 were detected in the plasma fraction of d < 1.006 g/mL of the PHT rabbit. The postprandial increase in apolipoprotein B in the PHT rabbit reflects a numerical increase in lipoprotein particles in the blood; the increase in apolipoproteins C-II and E suggests some disturbance in lipoprotein catabolism. Apolipoprotein B48 was detected postprandially in PHT rabbits. These results suggest that delayed catabolism of exogenous lipids caused the retention of chylomicron remnants in the blood. Results also suggest that activities of the lipolytic enzyme lipoprotein lipase and hepatic triglyceride lipase were deficient and that the hepatic uptake of exogenous lipoproteins was delayed in the PHT rabbit. Especially, for examining remnant hyperlipoproteinaemia in humans, PHT rabbit is an excellent animal model for hypertriglyceridaemia research.

  8. Apolipoprotein A-I in Labeo rohita: Cloning and functional characterisation reveal its broad spectrum antimicrobial property, and indicate significant role during ectoparasitic infection.

    PubMed

    Mohapatra, Amruta; Karan, Sweta; Kar, Banya; Garg, L C; Dixit, A; Sahoo, P K

    2016-08-01

    Apolipoprotein A-I (ApoA-I) is the most abundant and multifunctional high-density lipoprotein (HDL) having a major role in lipid transport and potent antimicrobial activity against a wide range of microbes. In this study, a complete CDS of 771 bp of Labeo rohita (rohu) ApoA-I (LrApoA-I) encoding a protein of 256 amino acids was amplified, cloned and sequenced. Tissue specific transcription analysis of LrApoA-I revealed its expression in a wide range of tissues, with a very high level of expression in liver and spleen. Ontogenic study of LrApoA-I expression showed presence of transcripts in milt and 3 h post-fertilization onwards in the larvae. The expression kinetics of LrApoA-I was studied upon infection with three different types of pathogens to elucidate its functional significance. Its expression was found to be up-regulated in the anterior kidney of L. rohita post-infection with Aeromonas hydrophila. Similarly following poly I:C (poly inosinic:cytidylic) stimulation, the transcript levels increased in both the anterior kidney and liver tissues. Significant up-regulation of LrApoA-I expression was observed in skin, mucous, liver and anterior kidney of the fish challenged with the ectoparasite Argulus siamensis. Immunomodulatory effect of recombinant LrApoA-I (rApoA-I) produced in Escherichia coli was demonstrated against A. hydrophila challenge in vivo. L. rohita administered with rApoA-I at a dose of 100 μg exhibited significantly higher protection (∼55%) upon challenge with A. hydrophila 12 h post-administration of the protein, in comparison to that observed in control group, along with higher level of expression of immune-related genes. The heightened expression of ApoA-I observed post-infection reflected its involvement in immune responses against a wide range of infections including bacterial, viral as well as parasitic pathogens. Our results also suggest the possibility of using rApoA-I as an immunostimulant, particularly rendering protection

  9. Quantification of serum apolipoproteins A-I and B-100 in clinical samples using an automated SISCAPA-MALDI-TOF-MS workflow.

    PubMed

    van den Broek, Irene; Nouta, Jan; Razavi, Morteza; Yip, Richard; Bladergroen, Marco R; Romijn, Fred P H T M; Smit, Nico P M; Drews, Oliver; Paape, Rainer; Suckau, Detlev; Deelder, André M; van der Burgt, Yuri E M; Pearson, Terry W; Anderson, N Leigh; Cobbaert, Christa M

    2015-06-15

    A fully automated workflow was developed and validated for simultaneous quantification of the cardiovascular disease risk markers apolipoproteins A-I (apoA-I) and B-100 (apoB-100) in clinical sera. By coupling of stable-isotope standards and capture by anti-peptide antibodies (SISCAPA) for enrichment of proteotypic peptides from serum digests to matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS detection, the standardized platform enabled rapid, liquid chromatography-free quantification at a relatively high throughput of 96 samples in 12h. The average imprecision in normo- and triglyceridemic serum pools was 3.8% for apoA-I and 4.2% for apoB-100 (4 replicates over 5 days). If stored properly, the MALDI target containing enriched apoA-1 and apoB-100 peptides could be re-analyzed without any effect on bias or imprecision for at least 7 days after initial analysis. Validation of the workflow revealed excellent linearity for daily calibration with external, serum-based calibrators (R(2) of 0.984 for apoA-I and 0.976 for apoB-100 as average over five days), and absence of matrix effects or interference from triglycerides, protein content, hemolysates, or bilirubins. Quantification of apoA-I in 93 normo- and hypertriglyceridemic clinical sera showed good agreement with immunoturbidimetric analysis (slope = 1.01, R(2) = 0.95, mean bias = 4.0%). Measurement of apoB-100 in the same clinical sera using both methods, however, revealed several outliers in SISCAPA-MALDI-TOF-MS measurements, possibly as a result of the lower MALDI-TOF-MS signal intensity (slope = 1.09, R(2) = 0.91, mean bias = 2.0%). The combination of analytical performance, rapid cycle time and automation potential validate the SISCAPA-MALDI-TOF-MS platform as a valuable approach for standardized and high-throughput quantification of apoA-I and apoB-100 in large sample cohorts. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Amyloid-Forming Properties of Human Apolipoproteins: Sequence Analyses and Structural Insights

    PubMed Central

    Das, Madhurima

    2017-01-01

    Apolipoproteins are protein constituents of lipoproteins that transport cholesterol and fat in circulation and are central to cardiovascular health and disease. Soluble apolipoproteins can transiently dissociate from the lipoprotein surface in a labile free form that can misfold, potentially leading to amyloid disease. Misfolding of apoA-I, apoA-II, and serum amyloid A (SAA) causes systemic amyloidoses, apoE4 is a critical risk factor in Alzheimer’s disease, and apolipoprotein misfolding is also implicated in cardiovascular disease. To explain why apolipoproteins are over- represented in amyloidoses, it was proposed that the amphipathic α-helices, which form the lipid surface-binding motif in this protein family, have high amyloid-forming propensity. Here, we use 12 sequence-based bioinformatics approaches to assess amyloid-forming potential of human apolipoproteins and to identify segments that are likely to initiate β-aggregation. Mapping such segments on the available atomic structures of apolipoproteins helps explain why some of them readily form amyloid while others do not. Our analysis shows that nearly all amyloidogenic segments: (i) are largely hydrophobic, (ii) are located in the lipid-binding amphipathic α-helices in the native structures of soluble apolipoproteins, (iii) are predicted in both native α-helices and β-sheets in the insoluble apoB, and (iv) are predicted to form parallel in-register β-sheet in amyloid. Most of these predictions have been verified experimentally for apoC-II, apoA-I, apoA-II and SAA. Surprisingly, the rank order of the amino acid sequence propensity to form amyloid (apoB > apoA-II > apoC-II ≥ apoA-I, apoC-III, SAA, apoC-I > apoA-IV, apoA-V, apoE) does not correlate with the proteins’ involvement in amyloidosis. Rather, it correlates directly with the strength of the protein-lipid association, which increases with increasing protein hydrophobicity. Therefore, the lipid surface-binding function and the amyloid

  11. Human Cathelicidin Compensates for the Role of Apolipoproteins in Hepatitis C Virus Infectious Particle Formation

    PubMed Central

    Puig-Basagoiti, Francesc; Fukuhara, Takasuke; Tamura, Tomokazu; Ono, Chikako; Uemura, Kentaro; Kawachi, Yukako; Yamamoto, Satomi; Mori, Hiroyuki; Kurihara, Takeshi; Okamoto, Toru; Aizaki, Hideki

    2016-01-01

    ABSTRACT Exchangeable apolipoproteins (ApoA, -C, and -E) have been shown to redundantly participate in the formation of infectious hepatitis C virus (HCV) particles during the assembly process, although their precise role in the viral life cycle is not well understood. Recently, it was shown that the exogenous expression of only short sequences containing amphipathic α-helices from various apolipoproteins is sufficient to restore the formation of infectious HCV particles in ApoB and ApoE double-gene-knockout Huh7 (BE-KO) cells. In this study, through the expression of a small library of human secretory proteins containing amphipathic α-helix structures, we identified the human cathelicidin antimicrobial peptide (CAMP), the only known member of the cathelicidin family of antimicrobial peptides (AMPs) in humans and expressed mainly in bone marrow and leukocytes. We showed that CAMP is able to rescue HCV infectious particle formation in BE-KO cells. In addition, we revealed that the LL-37 domain in CAMP containing amphipathic α-helices is crucial for the compensation of infectivity in BE-KO cells, and the expression of CAMP in nonhepatic 293T cells expressing claudin 1 and microRNA miR-122 confers complete propagation of HCV. These results suggest the possibility of extrahepatic propagation of HCV in cells with low-level or no expression of apolipoproteins but expressing secretory proteins containing amphipathic α-helices such as CAMP. IMPORTANCE Various exchangeable apolipoproteins play a pivotal role in the formation of infectious HCV during the assembly of viral particles, and amphipathic α-helix motifs in the apolipoproteins have been shown to be a key factor. To the best of our knowledge, we have identified for the first time the human cathelicidin CAMP as a cellular protein that can compensate for the role of apolipoproteins in the life cycle of HCV. We have also identified the domain in CAMP that contains amphipathic α-helices crucial for compensation and

  12. Influence of domain stability on the properties of human apolipoprotein E3 and E4 and mouse apolipoprotein E.

    PubMed

    Nguyen, David; Dhanasekaran, Padmaja; Nickel, Margaret; Mizuguchi, Chiharu; Watanabe, Mayu; Saito, Hiroyuki; Phillips, Michael C; Lund-Katz, Sissel

    2014-06-24

    The human apolipoprotein (apo) E4 isoform, which differs from wild-type apoE3 by the single amino acid substitution C112R, is associated with elevated risk of cardiovascular and Alzheimer’s diseases, but the molecular basis for this variation between isoforms is not understood. Human apoE is a two-domain protein comprising an N-terminal helix bundle and a separately folded C-terminal region. Here, we examine the concept that the ability of the protein to bind to lipid surfaces is influenced by the stability (or readiness to unfold) of these domains. The lipid-free structures and abilities to bind to lipid and lipoprotein particles of a series of human and mouse apoE variants with varying domain stabilities and domain–domain interactions are compared. As assessed by urea denaturation, the two domains are more unstable in apoE4 than in apoE3. To distinguish the contributions of the destabilization of each domain to the greater lipid-binding ability of apoE4, the properties of the apoE4 R61T and E255A variants, which have the same helix bundle stabilities but altered C-terminal domain stabilities, are compared. In these cases, the effects on lipid-binding properties are relatively minor, indicating that the destabilization of the helix bundle domain is primarily responsible for the enhanced lipid-binding ability of apoE4. Unlike human apoE, mouse apoE behaves essentially as a single domain, and its lipid-binding characteristics are more similar to those of apoE4. Together, the results show that the overall stability of the entire apoE molecule exerts a major influence on its lipid- and lipoprotein-binding properties.

  13. Lipoprotein remodeling generates lipid-poor apolipoprotein A-I particles in human interstitial fluid

    PubMed Central

    Olszewski, Waldemar L.; Hattori, Hiroaki; Miller, Irina P.; Kujiraoka, Takeshi; Oka, Tomoichiro; Iwasaki, Tadao; Nanjee, M. Nazeem

    2013-01-01

    Although much is known about the remodeling of high density lipoproteins (HDLs) in blood, there is no information on that in interstitial fluid, where it might have a major impact on the transport of cholesterol from cells. We incubated plasma and afferent (prenodal) peripheral lymph from 10 healthy men at 37°C in vitro and followed the changes in HDL subclasses by nondenaturing two-dimensional crossed immunoelectrophoresis and size-exclusion chromatography. In plasma, there was always initially a net conversion of small pre-β-HDLs to cholesteryl ester (CE)-rich α-HDLs. By contrast, in lymph, there was only net production of pre-β-HDLs from α-HDLs. Endogenous cholesterol esterification rate, cholesteryl ester transfer protein (CETP) concentration, CE transfer activity, phospholipid transfer protein (PLTP) concentration, and phospholipid transfer activity in lymph averaged 5.0, 10.4, 8.2, 25.0, and 82.0% of those in plasma, respectively (all P < 0.02). Lymph PLTP concentration, but not phospholipid transfer activity, was positively correlated with that in plasma (r = +0.63, P = 0.05). Mean PLTP-specific activity was 3.5-fold greater in lymph, reflecting a greater proportion of the high-activity form of PLTP. These findings suggest that cholesterol esterification rate and PLTP specific activity are differentially regulated in the two matrices in accordance with the requirements of reverse cholesterol transport, generating lipid-poor pre-β-HDLs in the extracellular matrix for cholesterol uptake from neighboring cells and converting pre-β-HDLs to α-HDLs in plasma for the delivery of cell-derived CEs to the liver. PMID:23233540

  14. Apolipoprotein modulation of streptococcal serum opacity factor activity against human plasma high-density lipoproteins.

    PubMed

    Rosales, Corina; Gillard, Baiba K; Courtney, Harry S; Blanco-Vaca, Francisco; Pownall, Henry J

    2009-08-25

    Human plasma HDL are the target of streptococcal serum opacity factor (SOF), a virulence factor that clouds human plasma. Recombinant (r) SOF transfers cholesteryl esters (CE) from approximately 400,000 HDL particles to a CE-rich microemulsion (CERM), forms a cholesterol-poor HDL-like particle (neo HDL), and releases lipid-free (LF) apo A-I. Whereas the rSOF reaction requires labile apo A-I, the modulation effects of other apos are not known. We compared the products and rates of the rSOF reaction against human HDL and HDL from mice overexpressing apos A-I and A-II. Kinetic studies showed that the reactivity of various HDL species is apo-specific. LpA-I reacts faster than LpA-I/A-II. Adding apos A-I and A-II inhibited the SOF reaction, an effect that was more profound for apo A-II. The rate of SOF-mediated CERM formation was slower against HDL from mice expressing human apos A-I and A-II than against WT mice HDL and slowest against HDL from apo A-II overexpressing mice. The lower reactivity of SOF against HDL containing human apos is due to the higher hydropathy of human apo A-I, particularly its C-terminus relative to mouse apo A-I, and the higher lipophilicity of human apo A-II. The SOF-catalyzed reaction is the first to target HDL rather than its transporters and receptors in a way that enhances reverse cholesterol transport (RCT). Thus, effects of apos on the SOF reaction are highly relevant. Our studies show that the "humanized" apo A-I-expressing mouse is a good animal model for studies of rSOF effects on RCT in vivo.

  15. Activation of lecithin cholesterol acyltransferase by human apolipoprotein E in discoidal complexes with lipids.

    PubMed

    Zorich, N; Jonas, A; Pownall, H J

    1985-07-25

    In a continued investigation of lecithin cholesterol acyltransferase reaction with micellar discoidal complexes of phosphatidylcholine, cholesterol, and various water soluble apolipoproteins, we prepared complexes containing human apo-E by the cholate dialysis method. These complexes were systematically compared to apo-A-I complexes synthesized under the same reaction conditions. Apo-E complexes (134 A in diameter) were slightly larger than apo-A-I complexes (110 A) but were very similar in terms of their protein and lipid content (2.4:0.10:1.0, egg phosphatidylcholine/cholesterol/apolipoprotein, w/w) and in the percentage of apolipoprotein in alpha-helical structure (72-74%). Concentration and temperature-dependence experiments on the velocity of the lecithin cholesterol acyltransferase reaction revealed differences in apparent Km values and small differences in apparent Vmax but very similar activation energies (18-20 kcal/mol). These observations suggest that differences in lecithin cholesterol acyltransferase activation by apo-A-I and apo-E are primarily a result of different affinities of the enzyme for the particles but that the rate-limiting step of the reaction is comparable for both complexes. Apo-E was found to be 18% as effective as apo-A-I in activating purified human lecithin cholesterol acyltransferase. Addition of free apo-A-I to apo-E complexes resulted in the exchange of bound for free apolipoprotein causing a slight increase in the reactivity with the enzyme when the incubation mixture was assayed. When the unbound apolipoproteins were removed by ultracentrifugation reisolated complexes containing both apo-E and apo-A-I demonstrated an even greater increase in reactivity with the enzyme.

  16. Gene-centric Association Signals for Lipids and Apolipoproteins Identified via the HumanCVD BeadChip

    PubMed Central

    Talmud, Philippa J.; Drenos, Fotios; Shah, Sonia; Shah, Tina; Palmen, Jutta; Verzilli, Claudio; Gaunt, Tom R.; Pallas, Jacky; Lovering, Ruth; Li, Kawah; Casas, Juan Pablo; Sofat, Reecha; Kumari, Meena; Rodriguez, Santiago; Johnson, Toby; Newhouse, Stephen J.; Dominiczak, Anna; Samani, Nilesh J.; Caulfield, Mark; Sever, Peter; Stanton, Alice; Shields, Denis C.; Padmanabhan, Sandosh; Melander, Olle; Hastie, Claire; Delles, Christian; Ebrahim, Shah; Marmot, Michael G.; Smith, George Davey; Lawlor, Debbie A.; Munroe, Patricia B.; Day, Ian N.; Kivimaki, Mika; Whittaker, John; Humphries, Steve E.; Hingorani, Aroon D.

    2009-01-01

    Blood lipids are important cardiovascular disease (CVD) risk factors with both genetic and environmental determinants. The Whitehall II study (n = 5592) was genotyped with the gene-centric HumanCVD BeadChip (Illumina). We identified 195 SNPs in 16 genes/regions associated with 3 major lipid fractions and 2 apolipoprotein components at p < 10−5, with the associations being broadly concordant with prior genome-wide analysis. SNPs associated with LDL cholesterol and apolipoprotein B were located in LDLR, PCSK9, APOB, CELSR2, HMGCR, CETP, the TOMM40-APOE-C1-C2-C4 cluster, and the APOA5-A4-C3-A1 cluster; SNPs associated with HDL cholesterol and apolipoprotein AI were in CETP, LPL, LIPC, APOA5-A4-C3-A1, and ABCA1; and SNPs associated with triglycerides in GCKR, BAZ1B, MLXIPL, LPL, and APOA5-A4-C3-A1. For 48 SNPs in previously unreported loci that were significant at p < 10−4 in Whitehall II, in silico analysis including the British Women's Heart and Health Study, BRIGHT, ASCOT, and NORDIL studies (total n > 12,500) revealed previously unreported associations of SH2B3 (p < 2.2 × 10−6), BMPR2 (p < 2.3 × 10−7), BCL3/PVRL2 (flanking APOE; p < 4.4 × 10−8), and SMARCA4 (flanking LDLR; p < 2.5 × 10−7) with LDL cholesterol. Common alleles in these genes explained 6.1%–14.7% of the variance in the five lipid-related traits, and individuals at opposite tails of the additive allele score exhibited substantial differences in trait levels (e.g., >1 mmol/L in LDL cholesterol [∼1 SD of the trait distribution]). These data suggest that multiple common alleles of small effect can make important contributions to individual differences in blood lipids potentially relevant to the assessment of CVD risk. These genes provide further insights into lipid metabolism and the likely effects of modifying the encoded targets therapeutically. PMID:19913121

  17. Lipoprotein lipase, LDL receptors and apo-lipoproteins in human fetal membranes at term.

    PubMed

    Huter, O; Wolf, H J; Schnetzer, A; Pfaller, K

    1997-11-01

    Ultrastructurally, all cells of human fetal membranes strongly exhibit a large amount of lipid deposits throughout pregnancy. Their origin and function is still unknown. The aim of this study was to investigate the localization of key components of lipid metabolism in this tissue. Using immunohistochemical techniques, the distribution of lipoprotein lipase (LPL), low density lipoprotein receptors (LDL receptors), and apo-lipoprotein B and E was investigated in 20 human fetal membranes at term. In addition, electron microscopy was used to study the intracellular localization of lipoprotein-sized particles. Amnionic epithelium and trophoblast cells reacted strongly for LPL. LDL receptors and apo-lipoproteins were present in amnionic epithelium and fibroblasts of the amnion. In none of the investigated cells were lipoprotein-sized particles identified. Similar results were obtained in all 20 cases. The findings indicate that lipoprotein from the amniotic fluid or from the maternal circulation may serve as substrate for lipids in human fetal membranes.

  18. Human liver apolipoprotein B-100 cDNA: complete nucleic acid and derived amino acid sequence.

    PubMed Central

    Law, S W; Grant, S M; Higuchi, K; Hospattankar, A; Lackner, K; Lee, N; Brewer, H B

    1986-01-01

    Human apolipoprotein B-100 (apoB-100), the ligand on low density lipoproteins that interacts with the low density lipoprotein receptor and initiates receptor-mediated endocytosis and low density lipoprotein catabolism, has been cloned, and the complete nucleic acid and derived amino acid sequences have been determined. ApoB-100 cDNAs were isolated from normal human liver cDNA libraries utilizing immunoscreening as well as filter hybridization with radiolabeled apoB-100 oligodeoxynucleotides. The apoB-100 mRNA is 14.1 kilobases long encoding a mature apoB-100 protein of 4536 amino acids with a calculated amino acid molecular weight of 512,723. ApoB-100 contains 20 potential glycosylation sites, and 12 of a total of 25 cysteine residues are located in the amino-terminal region of the apolipoprotein providing a potential globular structure of the amino terminus of the protein. ApoB-100 contains relatively few regions of amphipathic helices, but compared to other human apolipoproteins it is enriched in beta-structure. The delineation of the entire human apoB-100 sequence will now permit a detailed analysis of the conformation of the protein, the low density lipoprotein receptor binding domain(s), and the structural relationship between apoB-100 and apoB-48 and will provide the basis for the study of genetic defects in apoB-100 in patients with dyslipoproteinemias. PMID:3464946

  19. Several Human Liver Cell Expressed Apolipoproteins Complement HCV Virus Production with Varying Efficacy Conferring Differential Specific Infectivity to Released Viruses

    PubMed Central

    Doepke, Mandy; Vieyres, Gabrielle; Todt, Daniel; Wölk, Benno; Vondran, Florian W. R.; Geffers, Robert; Lauber, Chris; Kaderali, Lars; Penin, François; Pietschmann, Thomas

    2015-01-01

    Apolipoprotein E (ApoE), an exchangeable apolipoprotein, is necessary for production of infectious Hepatitis C virus (HCV) particles. However, ApoE is not the only liver-expressed apolipoprotein and the role of other apolipoproteins for production of infectious HCV progeny is incompletely defined. Therefore, we quantified mRNA expression of human apolipoproteins in primary human hepatocytes. Subsequently, cDNAs encoding apolipoproteins were expressed in 293T/miR-122 cells to explore if they complement HCV virus production in cells that are non-permissive due to limiting endogenous levels of human apolipoproteins. Primary human hepatocytes expressed high mRNA levels of ApoA1, A2, C1, C3, E, and H. ApoA4, A5, B, D, F, J, L1, L2, L3, L4, L6, M, and O were expressed at intermediate levels, and C2, C4, and L5 were not detected. All members of the ApoA and ApoC family of lipoproteins complemented HCV virus production in HCV transfected 293T/miR-122 cells, albeit with significantly lower efficacy compared with ApoE. In contrast, ApoD expression did not support production of infectious HCV. Specific infectivity of released particles complemented with ApoA family members was significantly lower compared with ApoE. Moreover, the ratio of extracellular to intracellular infectious virus was significantly higher for ApoE compared to ApoA2 and ApoC3. Since apolipoproteins complementing HCV virus production share amphipathic alpha helices as common structural features we altered the two alpha helices of ApoC1. Helix breaking mutations in both ApoC1 helices impaired virus assembly highlighting a critical role of alpha helices in apolipoproteins supporting HCV assembly. In summary, various liver expressed apolipoproteins with amphipathic alpha helices complement HCV virus production in human non liver cells. Differences in the efficiency of virus assembly, the specific infectivity of released particles, and the ratio between extracellular and intracellular infectivity point to

  20. Lipid-Free Apolipoprotein A-I Reduces Progression of Atherosclerosis by Mobilizing Microdomain Cholesterol and Attenuating the Number of CD131 Expressing Cells: Monitoring Cholesterol Homeostasis Using the Cellular Ester to Total Cholesterol Ratio.

    PubMed

    Kaul, Sushma; Xu, Hao; Zabalawi, Manal; Maruko, Elisa; Fulp, Brian E; Bluemn, Theresa; Brzoza-Lewis, Kristina L; Gerelus, Mark; Weerasekera, Ranjuna; Kallinger, Rachel; James, Roland; Zhang, Yi Sherry; Thomas, Michael J; Sorci-Thomas, Mary G

    2016-11-07

    Atherosclerosis is a chronic inflammatory disorder whose development is inversely correlated with high-density lipoprotein concentration. Current therapies involve pharmaceuticals that significantly elevate plasma high-density lipoprotein cholesterol concentrations. Our studies were conducted to investigate the effects of low-dose lipid-free apolipoprotein A-I (apoA-I) on chronic inflammation. The aims of these studies were to determine how subcutaneously injected lipid-free apoA-I reduces accumulation of lipid and immune cells within the aortic root of hypercholesterolemic mice without sustained elevations in plasma high-density lipoprotein cholesterol concentrations. Ldlr(-/-) and Ldlr(-/-) apoA-I(-/-) mice were fed a Western diet for a total of 12 weeks. After 6 weeks, a subset of mice from each group received subcutaneous injections of 200 μg of lipid-free human apoA-I 3 times a week, while the other subset received 200 μg of albumin, as a control. Mice treated with lipid-free apoA-I showed a decrease in cholesterol deposition and immune cell retention in the aortic root compared with albumin-treated mice, regardless of genotype. This reduction in atherosclerosis appeared to be directly related to a decrease in the number of CD131 expressing cells and the esterified cholesterol to total cholesterol content in several immune cell compartments. In addition, apoA-I treatment altered microdomain cholesterol composition that shifted CD131, the common β subunit of the interleukin 3 receptor, from lipid raft to nonraft fractions of the plasma membrane. ApoA-I treatment reduced lipid and immune cell accumulation within the aortic root by systemically reducing microdomain cholesterol content in immune cells. These data suggest that lipid-free apoA-I mediates beneficial effects through attenuation of immune cell lipid raft cholesterol content, which affects numerous types of signal transduction pathways that rely on microdomain integrity for assembly and

  1. Safety and Tolerability of CSL112, a Reconstituted, Infusible, Plasma-Derived Apolipoprotein A-I, After Acute Myocardial Infarction: The AEGIS-I Trial (ApoA-I Event Reducing in Ischemic Syndromes I).

    PubMed

    Michael Gibson, C; Korjian, Serge; Tricoci, Pierluigi; Daaboul, Yazan; Yee, Megan; Jain, Purva; Alexander, John H; Steg, P Gabriel; Lincoff, A Michael; Kastelein, John J P; Mehran, Roxana; D'Andrea, Denise M; Deckelbaum, Lawrence I; Merkely, Bela; Zarebinski, Maciej; Ophuis, Ton Oude; Harrington, Robert A

    2016-12-13

    Human or recombinant apolipoprotein A-I (apoA-I) has been shown to increase high-density lipoprotein-mediated cholesterol efflux capacity and to regress atherosclerotic disease in animal and clinical studies. CSL112 is an infusible, plasma-derived apoA-I that has been studied in normal subjects or those with stable coronary artery disease. This study aimed to characterize the safety, tolerability, pharmacokinetics, and pharmacodynamics of CSL112 in patients with a recent acute myocardial infarction. The AEGIS-I trial (Apo-I Event Reducing in Ischemic Syndromes I) was a multicenter, randomized, double-blind, placebo-controlled, dose-ranging phase 2b trial. Patients with myocardial infarction were stratified by renal function and randomized 1:1:1 to CSL112 (2 g apoA-I per dose) and high-dose CSL112 (6 g apoA-I per dose), or placebo for 4 consecutive weekly infusions. Coprimary safety end points were occurrence of either a hepatic safety event (an increase in alanine transaminase >3 times the upper limit of normal or an increase in total bilirubin >2 times the upper limit of normal) or a renal safety event (an increase in serum creatinine >1.5 times the baseline value or a new requirement for renal replacement therapy). A total of 1258 patients were randomized, and 91.2% received all 4 infusions. The difference in incidence rates for an increase in alanine transaminase or total bilirubin between both CSL112 arms and placebo was within the protocol-defined noninferiority margin of 4%. Similarly, the difference in incidence rates for an increase in serum creatinine or a new requirement for renal replacement therapy was within the protocol-defined noninferiority margin of 5%. CSL112 was associated with increases in apoA-I and ex vivo cholesterol efflux similar to that achieved in patients with stable coronary artery disease. In regard to the secondary efficacy end point, the risk for the composite of major adverse cardiovascular events among the groups was similar. Among

  2. Expression of human apolipoprotein B and assembly of lipoprotein(a) in transgenic mice

    SciTech Connect

    Callow, M.J.; Stoltzfus, L.J.; Rubin, E.M.; Lawn, R.M.

    1994-03-15

    The atherogenic macromolecule lipoprotein(a) [Lp(a)] has resisted in vivo analyses partly because it is found in a limited number of experimental animals. Although transgenic mice expressing human apolipoprotein (a) [apo(a)] have previously been described, they failed to assemble Lp(a) particles because of the inability of human apo(a) to associate with mouse apolipoprotein B (apoB). The authors isolated a 90-kilobase P1 phagemid containing the human apoB gene and with this DNA generated 13 lines of transgenic mice of which 11 expressed human apoB. The human apoB transcript was expressed and edited in the liver of the transgenic mice. Plasma concentrations of human apoB, as well as low density lipoprotein (LDL), were related to transgene copy number; the transgenic line with the most copies of human apoB had a >4-fold increase in LDL cholesterol compared with nontransgenics and a lipoprotein profile similar to that of humans. When human apoB and apo(a) transgenic mice were bred together, plasma apo(a) in mice expressing both human proteins was tightly associated with lipoproteins in the LDL density region. These studies demonstrate the successful expression of human apoB and the efficient assembly of Lp(a) in mice.

  3. Characterization of monoclonal antibodies against human apolipoprotein E.

    PubMed Central

    Milne, R W; Douste-Blazy, P; Marcel, Y L; Retegui, L

    1981-01-01

    From a single cell fusion, five stable hybridomas secreting antiapolipoprotein E (apo E) were obtained. The immunoglobulin (Ig)G subclasses containing the respective monoclonal antibodies were isolated and were used as the antibody component in a solid-phase radioimmunoassay. The binding of 125I-apo E to the insolubilized antibody was inhibited by unlabeled apo E but not by unlabeled apoproteins A-I, A-II, C-II, and C-III, or by low density lipoprotein immunodepleted of endogenous apo E. Competition curves were obtained with lipoprotein subfractions that had the same shape as those obtained with purified apo E. Apo E levels in normal and hyperlipidemic plasma were well correlated when measured by the five monoclonal antibodies and polyclonal anti-apo E, although differences in absolute values were observed. In normal subjects 34, 10, 20, and 36% of apo E was recovered in the very low density lipoprotein, low density lipoprotein, high density lipoprotein, and the d greater than 1.21-gl/ml fractions, respectively, whereas these values were 34, 7, 12, and 47%, respectively, in type III patients. All antibodies indicated the same subfraction distribution of apo E. The monoclonal antibodies reacted with all of the isomorphs of apo E. One of the antibodies could be clearly distinguished by its reactivity with chemically modified very low density lipoprotein. Images PMID:6788802

  4. Serum amyloid A-containing human high density lipoprotein 3. Density, size, and apolipoprotein composition.

    PubMed

    Coetzee, G A; Strachan, A F; van der Westhuyzen, D R; Hoppe, H C; Jeenah, M S; de Beer, F C

    1986-07-25

    Serum amyloid A protein (apo-SAA), an acute phase reactant, is an apolipoprotein of high density lipoproteins (HDL), in particular the denser subpopulation HDL3. The structure of HDL3 isolated from humans affected by a variety of severe disease states was investigated with respect to density, size, and apolipoprotein composition, using density gradient ultracentrifugation, gradient gel electrophoresis, gel filtration, and solid phase immunoadsorption. Apo-SAA was present in HDL particles in increasing amounts as particle density increased. Apo-SAA-containing HDL3 had bigger radii than normal HDL3 of comparable density. Purified apo-SAA associated readily with normal HDL3 in vitro, giving rise to particles containing up to 80% of their apoproteins as apo-SAA. The addition of apo-SAA resulted in a displacement of apo-A-I and an increase in particle size. Acute phase HDL3 represented a mixture of particles, polydisperse with respect to apolipoprotein content; for example, some particles were isolated that contained apo-A-I, apo-A-II, and apo-SAA, whereas others contained apo-A-I and apo-SAA but no apo-A-II. We conclude that apo-SAA probably associates in the circulation of acute phase patients with existing HDL particles, causing the remodeling of the HDL shell to yield particles of bigger size and higher density that are relatively depleted of apo-A-I.

  5. Glomerular Autoimmune Multicomponents of Human Lupus Nephritis In Vivo: α-Enolase and Annexin AI

    PubMed Central

    Bruschi, Maurizio; Sinico, Renato Alberto; Moroni, Gabriella; Pratesi, Federico; Migliorini, Paola; Galetti, Maricla; Murtas, Corrado; Tincani, Angela; Madaio, Michael; Radice, Antonella; Franceschini, Franco; Trezzi, Barbara; Bianchi, Laura; Giallongo, Agata; Gatti, Rita; Tardanico, Regina; Scaloni, Andrea; D’Ambrosio, Chiara; Carnevali, Maria Luisa; Messa, Piergiorgio; Ravani, Pietro; Barbano, Giancarlo; Bianco, Beatrice; Bonanni, Alice; Scolari, Francesco; Martini, Alberto; Candiano, Giovanni; Allegri, Landino

    2014-01-01

    Renal targets of autoimmunity in human lupus nephritis (LN) are unknown. We sought to identify autoantibodies and glomerular target antigens in renal biopsy samples from patients with LN and determine whether the same autoantibodies can be detected in circulation. Glomeruli were microdissected from biopsy samples of 20 patients with LN and characterized by proteomic techniques. Serum samples from large cohorts of patients with systemic lupus erythematosus (SLE) with and without LN and other glomerulonephritides were tested. Glomerular IgGs recognized 11 podocyte antigens, with reactivity varying by LN pathology. Notably, IgG2 autoantibodies against α-enolase and annexin AI were detected in 11 and 10 of the biopsy samples, respectively, and predominated over other autoantibodies. Immunohistochemistry revealed colocalization of α-enolase or annexin AI with IgG2 in glomeruli. High levels of serum anti–α-enolase (>15 mg/L) IgG2 and/or anti-annexin AI (>2.7 mg/L) IgG2 were detected in most patients with LN but not patients with other glomerulonephritides, and they identified two cohorts: patients with high anti–α-enolase/low anti-annexin AI IgG2 and patients with low anti–α-enolase/high anti-annexin AI IgG2. Serum levels of both autoantibodies decreased significantly after 12 months of therapy for LN. Anti–α-enolase IgG2 recognized specific epitopes of α-enolase and did not cross-react with dsDNA. Furthermore, nephritogenic monoclonal IgG2 (clone H147) derived from lupus-prone MRL-lpr/lpr mice recognized human α-enolase, suggesting homology between animal models and human LN. These data show a multiantibody composition in LN, where IgG2 autoantibodies against α-enolase and annexin AI predominate in the glomerulus and can be detected in serum. PMID:24790181

  6. Glomerular autoimmune multicomponents of human lupus nephritis in vivo: α-enolase and annexin AI.

    PubMed

    Bruschi, Maurizio; Sinico, Renato Alberto; Moroni, Gabriella; Pratesi, Federico; Migliorini, Paola; Galetti, Maricla; Murtas, Corrado; Tincani, Angela; Madaio, Michael; Radice, Antonella; Franceschini, Franco; Trezzi, Barbara; Bianchi, Laura; Giallongo, Agata; Gatti, Rita; Tardanico, Regina; Scaloni, Andrea; D'Ambrosio, Chiara; Carnevali, Maria Luisa; Messa, Piergiorgio; Ravani, Pietro; Barbano, Giancarlo; Bianco, Beatrice; Bonanni, Alice; Scolari, Francesco; Martini, Alberto; Candiano, Giovanni; Allegri, Landino; Ghiggeri, Gian Marco

    2014-11-01

    Renal targets of autoimmunity in human lupus nephritis (LN) are unknown. We sought to identify autoantibodies and glomerular target antigens in renal biopsy samples from patients with LN and determine whether the same autoantibodies can be detected in circulation. Glomeruli were microdissected from biopsy samples of 20 patients with LN and characterized by proteomic techniques. Serum samples from large cohorts of patients with systemic lupus erythematosus (SLE) with and without LN and other glomerulonephritides were tested. Glomerular IgGs recognized 11 podocyte antigens, with reactivity varying by LN pathology. Notably, IgG2 autoantibodies against α-enolase and annexin AI were detected in 11 and 10 of the biopsy samples, respectively, and predominated over other autoantibodies. Immunohistochemistry revealed colocalization of α-enolase or annexin AI with IgG2 in glomeruli. High levels of serum anti-α-enolase (>15 mg/L) IgG2 and/or anti-annexin AI (>2.7 mg/L) IgG2 were detected in most patients with LN but not patients with other glomerulonephritides, and they identified two cohorts: patients with high anti-α-enolase/low anti-annexin AI IgG2 and patients with low anti-α-enolase/high anti-annexin AI IgG2. Serum levels of both autoantibodies decreased significantly after 12 months of therapy for LN. Anti-α-enolase IgG2 recognized specific epitopes of α-enolase and did not cross-react with dsDNA. Furthermore, nephritogenic monoclonal IgG2 (clone H147) derived from lupus-prone MRL-lpr/lpr mice recognized human α-enolase, suggesting homology between animal models and human LN. These data show a multiantibody composition in LN, where IgG2 autoantibodies against α-enolase and annexin AI predominate in the glomerulus and can be detected in serum.

  7. Characterization of disulfide-linked heterodimers containing apolipoprotein D in human plasma lipoproteins.

    PubMed

    Blanco-Vaca, F; Via, D P; Yang, C Y; Massey, J B; Pownall, H J

    1992-12-01

    Human plasma apolipoprotein (apo) D is a glycoprotein with an apparent molecular weight of 29,000 M(r). It is present, mainly, in high density lipoproteins (HDL) and very high density lipoproteins (VHDL). Western blot analysis of HDL and VHDL using rabbit antibodies to human apoD revealed major immunoreactive bands at 29,000 and 38,000 M(r), with minor bands ranging from 50,000 to and 80,000 M(r). Only the 29,000 M(r) band corresponding to apoD remained when the electrophoresis was conducted under reducing conditions, demonstrating that apoD is cross-linked to other proteins via disulfide bonds. The broad pattern of immunoreactivity was also observed under nonreducing conditions when the blood was collected into a solution of sulfhydryl-trapping reagents, or when these reagents were added to the isolated lipoproteins. These results indicated that the disulfide bonds were not the result of disulfide exchange during the experimental procedures. On the basis of amino acid sequencing and reactions to antibodies, the 38,000 M(r) band was identified as an apoD-apoA-II heterodimer. The apoD-apoA-II was also demonstrated in plasma. In both HDL and plasma, the apoD-apoA-II heterodimer constituted the major form of apoD. Disulfide-linked heterodimers of apoD and apoB-100 were also found in low and very low density lipoproteins, and in whole plasma. It is concluded that a fraction of human apoD, like other cysteine-containing apolipoproteins, exists as a disulfide-linked heterodimer with other apolipoproteins in all major human lipoprotein fractions.

  8. Human leucine zipper protein promotes hepatic steatosis via induction of apolipoprotein A-IV.

    PubMed

    Kang, Minsoo; Kim, Jeonghan; An, Hyoung-Tae; Ko, Jesang

    2017-02-28

    The molecular mechanism of stress-induced hepatic steatosis is not well known. Human leucine zipper protein (LZIP) regulates the expression of genes involved in inflammation, cell migration, and stress response. The aim of this study was to determine the regulatory role of LZIP in stress-induced hepatic steatosis. We used a microarray analysis to identify LZIP-induced genes involved in hepatic lipid metabolism. LZIP increased the expression of apolipoprotein A-IV (APOA4) mRNA. In the presence of stress inducer, APOA4 promoter analysis was performed and LZIP-induced lipid accumulation was monitored in mouse primary cells and human tissues. Under Golgi stress conditions, LZIP underwent proteolytic cleavage and was phosphorylated by AKT to protect against proteasome degradation. The stabilized N-terminal LZIP was translocated to the nucleus, where it directly bound to the APOA4 promoter, leading to APOA4 induction. LZIP-induced APOA4 expression resulted in increased absorption of surrounding free fatty acids. LZIP also promoted hepatic steatosis in mouse liver. Both LZIP and APOA4 were highly expressed in human steatosis samples. Our findings indicate that LZIP is a novel modulator of APOA4 expression and hepatic lipid metabolism. LZIP might be a therapeutic target for developing treatment strategies for hepatic steatosis and related metabolic diseases.-Kang, M., Kim, J., An, H.-T., Ko, J. Human leucine zipper protein promotes hepatic steatosis via induction of apolipoprotein A-IV.

  9. Analysis of cell surface alterations in Legionella pneumophila cells treated with human apolipoprotein E.

    PubMed

    Palusinska-Szysz, Marta; Zdybicka-Barabas, Agnieszka; Cytryńska, Małgorzata; Wdowiak-Wróbel, Sylwia; Chmiel, Elżbieta; Gruszecki, Wiesław I

    2015-03-01

    Binding of human apolipoprotein E (apoE) to Legionella pneumophila lipopolysaccharide was analysed at the molecular level by Fourier-transform infrared spectroscopy, thereby providing biophysical evidence for apoE-L. pneumophila lipopolysaccharide interaction. Atomic force microscopy imaging of apoE-exposed L. pneumophila cells revealed alterations in the bacterial cell surface topography and nanomechanical properties in comparison with control bacteria. The changes induced by apoE binding to lipopolysaccharide on the surface of L. pneumophila cells may participate in: (1) impeding the penetration of host cells by the bacteria; (2) suppression of pathogen intracellular growth and eventually; and (3) inhibition of the development of infection.

  10. Human Apolipoprotein A-I-Derived Amyloid: Its Association with Atherosclerosis

    PubMed Central

    Ramella, Nahuel A.; Rimoldi, Omar J.; Prieto, Eduardo D.; Schinella, Guillermo R.; Sanchez, Susana A.; Jaureguiberry, María S.; Vela, María E.

    2011-01-01

    Amyloidoses constitute a group of diseases in which soluble proteins aggregate and deposit extracellularly in tissues. Nonhereditary apolipoprotein A-I (apoA-I) amyloid is characterized by deposits of nonvariant protein in atherosclerotic arteries. Despite being common, little is known about the pathogenesis and significance of apoA-I deposition. In this work we investigated by fluorescence and biochemical approaches the impact of a cellular microenvironment associated with chronic inflammation on the folding and pro-amyloidogenic processing of apoA-I. Results showed that mildly acidic pH promotes misfolding, aggregation, and increased binding of apoA-I to extracellular matrix elements, thus favoring protein deposition as amyloid like-complexes. In addition, activated neutrophils and oxidative/proteolytic cleavage of the protein give rise to pro amyloidogenic products. We conclude that, even though apoA-I is not inherently amyloidogenic, it may produce non hereditary amyloidosis as a consequence of the pro-inflammatory microenvironment associated to atherogenesis. PMID:21811627

  11. Myeloperoxidase-mediated oxidation targets serum apolipoprotein A-I in diabetic patients and represents a potential mechanism leading to impaired anti-apoptotic activity of high density lipoprotein.

    PubMed

    Lu, Naihao; Xie, Shiliang; Li, Jiayu; Tian, Rong; Peng, Yi-Yuan

    2015-02-20

    It is demonstrated that levels of protein-bound chlorotyrosine, nitrotyrosine and myeloperoxidase (MPO), a protein that catalyzes generation of chlorinating and nitrating oxidants, serve as independent predictors of cardiovascular disease. Immunoprecipitation and Western blot were used to analyze protein concentration, nitration and chlorination. LC-MS/MS was used to identify nitrated and chlorinated sites of Tyr from immunoprecipitated serum proteins. Apolipoprotein A-I (apoA-I), the primary protein constituent of high density lipoprotein (HDL), was identified as a selective target for MPO-catalyzed nitration and chlorination in patients with type 2 diabetes. The serum proteins from diabetic subjects showed that the levels of apoA-I nitration and chlorination were clearly increased, whereas apoA-I concentration and cholesterol efflux activity were significantly decreased. MPO as a likely mechanism for oxidative modification of apoA-I in vivo was apparently facilitated by MPO binding to apoA-I. Subsequently, it was found that Tyr 192 was the major nitration and chlorination site in apoA-I from diabetic serum. Further studies in vitro revealed that besides the classic inhibition in cholesterol efflux activities, MPO-catalyzed oxidation could result in a loss of anti-apoptotic activity of lipoprotein. ApoA-I undergoes MPO-mediated oxidation in serum from diabetic patients compared to non-diabetic subjects and MPO-catalyzed modification may impair the anti-apoptotic properties of HDL in vitro. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Dual Actions of Apolipoprotein A-I on Glucose-Stimulated Insulin Secretion and Insulin-Independent Peripheral Tissue Glucose Uptake Lead to Increased Heart and Skeletal Muscle Glucose Disposal.

    PubMed

    Domingo-Espín, Joan; Lindahl, Maria; Nilsson-Wolanin, Oktawia; Cushman, Samuel W; Stenkula, Karin G; Lagerstedt, Jens O

    2016-07-01

    Apolipoprotein A-I (apoA-I) of HDL is central to the transport of cholesterol in circulation. ApoA-I also provides glucose control with described in vitro effects of apoA-I on β-cell insulin secretion and muscle glucose uptake. In addition, apoA-I injections in insulin-resistant diet-induced obese (DIO) mice lead to increased glucose-stimulated insulin secretion (GSIS) and peripheral tissue glucose uptake. However, the relative contribution of apoA-I as an enhancer of GSIS in vivo and as a direct stimulator of insulin-independent glucose uptake is not known. Here, DIO mice with instant and transient blockade of insulin secretion were used in glucose tolerance tests and in positron emission tomography analyses. Data demonstrate that apoA-I to an equal extent enhances GSIS and acts as peripheral tissue activator of insulin-independent glucose uptake and verify skeletal muscle as an apoA-I target tissue. Intriguingly, our analyses also identify the heart as an important target tissue for the apoA-I-stimulated glucose uptake, with potential implications in diabetic cardiomyopathy. Explorations of apoA-I as a novel antidiabetic drug should extend to treatments of diabetic cardiomyopathy and other cardiovascular diseases in patients with diabetes. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  13. ABCA1 (ATP-Binding Cassette Transporter A1) Mediates ApoA-I (Apolipoprotein A-I) and ApoA-I Mimetic Peptide Mobilization of Extracellular Cholesterol Microdomains Deposited by Macrophages.

    PubMed

    Jin, Xueting; Sviridov, Denis; Liu, Ying; Vaisman, Boris; Addadi, Lia; Remaley, Alan T; Kruth, Howard S

    2016-12-01

    We examined the function of ABCA1 (ATP-binding cassette transporter A1) in ApoA-I (apolipoprotein A-I) mobilization of cholesterol microdomains deposited into the extracellular matrix by cholesterol-enriched macrophages. We have also determined whether an ApoA-I mimetic peptide without and with complexing to sphingomyelin can mobilize macrophage-deposited cholesterol microdomains. Extracellular cholesterol microdomains deposited by cholesterol-enriched macrophages were detected with a monoclonal antibody, 58B1. ApoA-I and an ApoA-I mimetic peptide 5A mobilized cholesterol microdomains deposited by ABCA1(+/+) macrophages but not by ABCA1(-/-) macrophages. In contrast, ApoA-I mimetic peptide 5A complexed with sphingomyelin could mobilize cholesterol microdomains deposited by ABCA1(-/-) macrophages. Our findings show that a unique pool of extracellular cholesterol microdomains deposited by macrophages can be mobilized by both ApoA-I and an ApoA-I mimetic peptide but that mobilization depends on macrophage ABCA1. It is known that ABCA1 complexes ApoA-I and ApoA-I mimetic peptide with phospholipid, a cholesterol-solubilizing agent, explaining the requirement for ABCA1 in extracellular cholesterol microdomain mobilization. Importantly, ApoA-I mimetic peptide already complexed with phospholipid can mobilize macrophage-deposited extracellular cholesterol microdomains even in the absence of ABCA1. © 2016 American Heart Association, Inc.

  14. Patterns of association between genetic variability in apolipoprotein (apo) B, apo AI-CIII-AIV, and cholesterol ester transfer protein gene regions and quantitative variation in lipid and lipoprotein traits: influence of gender and exogenous hormones.

    PubMed Central

    Kessling, A; Ouellette, S; Bouffard, O; Chamberland, A; Bétard, C; Selinger, E; Xhignesse, M; Lussier-Cacan, S; Davignon, J

    1992-01-01

    Patterns of RFLP association were studied, to identify gene regions influencing quantitative variation in lipid and lipoprotein traits (coronary artery disease [CAD] risk factors or metabolically related traits). Subjects (118 female and 229 male; age 20-59 years) were selected for health. Multiple RFLPs were used to sample variability in regions around genes for apolipoprotein (apo) B (restriction enzymes HincII, PvuII, EcoRI, and XbaI), apo AI-CIII-AIV (BamHI, XmnI, TaqI, PstI, SstI, and PvuII) and cholesterol ester transfer protein (TaqI). Separate analyses were done by gender. The sample was truncated at mean +/- 4 SD, to remove extreme outliers. There was no significant gender difference in RFLP genotype frequency distribution. After trait-level adjustment to maximize removal of concomitant variability, analysis of variance was used to estimate the percentage trait phenotypic variance explained by measured variability in the gene regions studied. Fewer gene regions were involved in men, with less influence on quantitative trait variation than in women, in whom hormone use affected association patterns. Gender differences imply that pooling genders or adjusting data for gender effects removes genetic information and should be avoided. The association patterns show that variability around the candidate genes modulates trait levels: the genes are contributors to the genetics of CAD risk variables in a healthy sample. PMID:1346081

  15. Controlling for apolipoprotein A-I concentrations changes the inverse direction of the relationship between high HDL-C concentration and a measure of pre-clinical atherosclerosis.

    PubMed

    Sung, Ki-Chul; Wild, Sarah H; Byrne, Christopher D

    2013-12-01

    The independent effect of high density lipoprotein cholesterol (HDL-C) concentration to confer cardiovascular disease protection has been questioned. We investigated whether the inverse association between HDL-C concentration and a measure of preclinical atherosclerosis was modified by other risk factors. Cross-sectional data were analysed from an occupational cohort of 12,031 men who had measurements of cardiovascular risk factors and a cardiac computed tomography (CT) estimation of coronary artery calcium (CAC) score, a measure of pre-clinical atherosclerosis. Logistic regression was used to describe associations between both HDL-C and Apo-A-I concentrations and their ratio as exposures, and CAC scores > 0, ≥ 20 and ≥ 100, as outcomes. 1351 (11.2%), 665 (5.5%) and 230 (1.9%) of participants had a CAC score > 0, ≥ 20 and ≥ 100, respectively. Adjusting for age, glucose, triglyceride, LDL-C, systolic blood pressure, waist circumference, prior cerebrovascular accident, prior coronary artery disease, prior hypertension, alcohol consumption, smoking status and exercise, a negative association existed between HDL-C and CAC score. (E.g. odds ratio (OR) for top compared to bottom HDL-C quartile for CAC > 0 = 0.78 [95%CI 0.64, 0.94], p = 0.01). Further adjustment for Apo A-I changed the direction of the association between HDL-C and CAC score > 0 (OR for top compared to bottom quartiles 1.61 [95%CI 1.18, 2.21], p = 0.003). Sensitivity analyses showed that point estimates for ORs were very similar regardless of CAC score threshold (CAC > 0, ≥ 20 and ≥ 100). Controlling for Apo A-I concentrations changes the inverse direction of relationship between high HDL-C concentration and a measure of pre-clinical atherosclerosis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  16. Docosahexaenoic acid prevents cognitive deficits in human apolipoprotein E epsilon 4-targeted replacement mice.

    PubMed

    Chouinard-Watkins, Raphaël; Vandal, Milène; Léveillé, Pauline; Pinçon, Anthony; Calon, Frédéric; Plourde, Mélanie

    2017-09-01

    At a population level, dietary consumption of fish rich in docosahexaenoic acid (DHA) is associated with prevention of cognitive decline but this association is not clear in carriers of the apolipoprotein E epsilon 4 allele (E4). Plasma and liver DHA concentrations show significant alterations in E4 carriers, in part corrected by DHA supplementation. However, whether DHA sufficiency in E4 carriers has consequences on cognition is unknown. Mice expressing human E4 or apolipoprotein E epsilon 3 allele (E3) were fed either a control diet or a diet containing DHA for 8 months and cognitive performance was tested using the object recognition test and the Barnes maze test. In E4 mice fed the control diet, impaired memory was detected and arachidonic acid concentrations were elevated in the hippocampus compared to E3 mice fed the control diet. DHA consumption prevented memory decline and restored arachidonic acid concentrations in the hippocampus of E4 mice. Our results suggest that long-term high-dose DHA intake may prevent cognitive decline in E4 carriers. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. [Isolation of isoforms of apolipoprotein CIII from human serum by chromatofocusing].

    PubMed

    Fang, D; Gong, R; O, K

    1999-03-01

    This study aimed to isolate isoforms of apolipoprotein (apo) C III from human serum. 24-hour fasting serum from normal and hyperlipidemic subjects was pooled and subjected to ultracentrifugation at plasma density for 20 hours. Very low density lipoprotein (VLDL) was collected at density of d < 1.006 g/ml, and it was delipidated by ethanol and ether. The delipidated apo-VLDL was dissolved in a solution containing 7.2 mol/L urea and 20 mmol/L dithiothreitol. The insoluble apo B was removed by centrifugation. The soluble apo-VLDL was applied to PBE94 column, and eluted with elution buffer containing polybuffer 74 and 8 mol/L urea (1:8, pH4.0). After pooled, the eluted peaks of apolipoproteins were applied to column chromatography of hydroxylapatite to remove the polybuffer. The purified isoforms of apoC III and the purified apo C I, C II and E, were characterized by isoelectrofocusing and west blot. The results showed that the purified apoC III1, C III2, and C II were pure.

  18. Transgenic mice expressing high plasma concentrations of human apolipoprotein B100 and lipoprotein(a).

    PubMed Central

    Linton, M F; Farese, R V; Chiesa, G; Grass, D S; Chin, P; Hammer, R E; Hobbs, H H; Young, S G

    1993-01-01

    The B apolipoproteins, apo-B48 and apo-B100, are key structural proteins in those classes of lipoproteins considered to be atherogenic [e.g., chylomicron remnants, beta-VLDL, LDL, oxidized LDL, and Lp(a)]. Here we describe the development of transgenic mice expressing high levels of human apo-B48 and apo-B100. A 79.5-kb human genomic DNA fragment containing the entire human apo-B gene was isolated from a P1 bacteriophage library and microinjected into fertilized mouse eggs. 16 transgenic founders expressing human apo-B were generated, and the animals with the highest expression had plasma apo-B100 levels nearly as high as those of normolipidemic humans (approximately 50 mg/dl). The human apo-B100 in transgenic mouse plasma was present largely in lipoproteins of the LDL class as shown by agarose gel electrophoresis, chromatography on a Superose 6 column, and density gradient ultracentrifugation. When the human apo-B transgenic founders were crossed with transgenic mice expressing human apo(a), the offspring that expressed both transgenes had high plasma levels of human Lp(a). Both the human apo-B and Lp(a) transgenic mice will be valuable resources for studying apo-B metabolism and the role of apo-B and Lp(a) in atherosclerosis. Images PMID:8254057

  19. Preparation of soluble apolipoproteins A-I, B, and C-II by a chromatofocusing column method, and evaluation of their concentrations in serum in pulmonary disease.

    PubMed

    Jauhiainen, M; Laitinen, M; Marniemi, J; Liippo, K; Penttilä, I; Hietanen, E

    1983-10-01

    A chromatofocusing column method for isolating ApoB is described. LDL is first isolated by sequential ultracentrifugation and delipidated with n-butanol/diisopropyl ether. Chromatofocusing of ApoLDL yielded a large ApoB peak at pI 5.0-5.3. ApoA-I and ApoC-II were prepared analogously, with HDL and VLDL as the source of apoprotein. Antisera were raised in rabbits, and electroimmunoassay techniques were used for determination. ApoB was water-soluble after chromatofocusing. Intra-assay precision (CV) was 4.7% for ApoA-I, 7.8% for ApoB in the "rocket" electrophoresis. Interassay precision (CV) was 6% for ApoA-I and 8% for ApoB. Apolipoprotein concentrations were measured in subjects who had undergone lung resection and patients with obstructive pulmonary disease. After lung resection, the concentration of ApoA-I in serum was significantly decreased (p less than 0.001) and that of ApoB significantly increased (p less than 0.001) as compared with controls. The ApoA-I/ApoB ratio was significantly lower in the lung-resection group. ApoA-I and ApoB concentrations were unchanged in chronic obstructive pulmonary disease. ApoC-II concentrations in each group were similar to those for control subjects. Of the lipids, values for total cholesterol were above normal after lung resection (p less than 0.002), as were those for triglycerides (p less than 0.02).

  20. Unusually high reactivity of apolipoprotein B-100 among proteins to radical reactions induced in human plasma.

    PubMed

    Hashimoto, R; Narita, S; Yamada, Y; Tanaka, K; Kojo, S

    2000-01-17

    Relative reactivities of proteins to radical reactions caused in human plasma were studied for the first time utilizing an immunoblotting assay. When radical reactions were caused by Cu(2+), apolipoprotein B-100 (apoB) underwent extensive fragmentation concurrently with the decrease in alpha-tocopherol, while human serum albumin (HSA) and transferrin (TF) were not decreased at all. When radical reactions were initiated by Cu(2+) with hydrogen peroxide or 2,2'-azobis(2-amidinopropane)dihydrochloride (AAPH), alpha-tocopherol and apoB were also decreased steadily but HSA and TF were not decreased. These observations indicate that apoB is extremely reactive, even comparable to alpha-tocopherol, towards radical reactions. These results also suggest that the radical reaction of apoB is a possible process in vivo and it is involved in atherogenesis along with low density lipoprotein lipid peroxidation, which has been studied extensively.

  1. Effect of Royal Jelly Intake on Serum Glucose, Apolipoprotein A-I (ApoA-I), Apolipoprotein B (ApoB) and ApoB/ApoA-I Ratios in Patients with Type 2 Diabetes: A Randomized, Double-Blind Clinical Trial Study.

    PubMed

    Khoshpey, Basemeh; Djazayeri, Shima; Amiri, Fatemehsadat; Malek, Mojtaba; Hosseini, Agha Fateme; Hosseini, Sharieh; Shidfar, Shahrzad; Shidfar, Farzad

    2016-08-01

    Type 2 diabetes is the most common metabolic disorder worldwide. Evidence supports a role for royal jelly (RJ) in reduction of serum glucose and lipids in animals and healthy subjects. The purpose of this study was to determine the effect of RJ intake on serum glucose, apolipoprotein A-I (ApoA-I), apolipoprotein B (ApoB) and ApoB/ApoA-I ratios in patients with type 2 diabetes. Fifty patients with type 2 diabetes participated in a double-blind, placebo-controlled study. The participants were randomly divided into RJ and placebo groups and were given doses of 1000 mg royal jelly or placebo 3 times a day for 8 weeks, respectively. Weight, height, fasting blood glucose, ApoA-I and ApoB were measured at baseline and endpoint. There were no significant differences in baseline characteristics and dietary intakes between groups. The mean difference in glucose concentrations decreased in the RJ group (-9.4 mg/dL vs. 4 mg/dL; p=0.011). The mean difference in ApoA-I concentrations increased in the RJ group (34.4 mg/dL vs. -1.08 mg/dL; p=0.013). There was a significant decrease in mean difference of ApoB/ApoA-I in the RJ group compared with the placebo group (0.008 vs. 0.13; p<0.044), respectively. These data suggest that RJ intake may have desirable effects on serum glucose, Apo-A-I concentrations and ApoB/ApoA-I ratios in people with type 2 diabetes. Copyright © 2016 Canadian Diabetes Association. Published by Elsevier Inc. All rights reserved.

  2. Orphan nuclear receptor Nur77 participates in human apolipoprotein A5 gene expression

    SciTech Connect

    Song, Kwang-Hoon

    2010-01-29

    The orphan nuclear receptor Nur77 (NR4A1) has been reported to play a crucial role in the modulation of diverse metabolic processes in liver. Here, we reported the identification of human apolipoprotein A5 (ApoA5), which implicated in lowering plasma triglyceride levels, as a novel target gene of Nur77. Nur77 induced the human ApoA5 promoter activity. Using 5'-deletion and mutagenesis of human ApoA5 promoter analysis and chromatin immunoprecipitation assays, it was shown that Nur77 directly regulated human ApoA5 gene expression by binding to a Nur77 response element (AAAGGTCA) located in the proximal human ApoA5 promoter region. In addition, we demonstrated that blocking of Nur77 transcriptional activity via overexpression of dominant negative Nur77 suppressed human ApoA5 promoter activity and mRNA expression in human hepatoma cells, HepG2. Taken together, our results demonstrated that Nur77 is a novel regulator of human ApoA5 gene expression and provide a new insight into the role of this orphan nuclear receptor in lipoprotein metabolism and triglyceride homeostasis.

  3. Quantitative measurement of lipoprotein particles containing both apolipoprotein AIV and apolipoprotein B in human plasma by a noncompetitive ELISA.

    PubMed

    Ferrer, Fanny; Bigot-Corbel, Edith; N'Guyen, Patrick; Krempf, Michel; Bard, Jean-Marie

    2002-06-01

    A reliable method for plasma would be useful to investigate the role of apolipoprotein (apo) AIV when associated with apo B-containing or triglyceride-rich lipoproteins. We used a sandwich ELISA to quantify lipoprotein B:AIV particles (Lp B:AIVf; lipoproteins containing at least apo B and apo AIV) in plasma. The method used microtiter plates coated with purified anti-apo B immunoglobulins that selectively retained apo B-containing particles. Lipoproteins containing both apo B and apo AIV were distinguished from those containing only apo B by use of a peroxidase-labeled anti-apo AIV antibody. These subspecies were revealed by ABTS reagent and further quantified by spectrophotometry. Results were expressed in mg/L apo AIV associated with apo B. This method was applied to samples with different cholesterol and triglyceride concentrations. The developed sandwich ELISA method identified and quantified Lp B:AIVf in plasma samples. Within- and between-run CVs were approximately 10%, and analytical recoveries were 95-107%. Results were not significantly influenced by addition of triglycerides or by storage at -20 degrees C (up to 9 months). Under these conditions, plasma Lp B:AIVf concentrations were statistically higher in hypercholesterolemic and mixed hyperlipidemic individuals (53 +/- 13 mg/L; P <0.001 and 70 +/- 18 mg/L; P <0.001, respectively) than in normolipidemic individuals (43 +/- 12 mg/L). Lp B:AIVf concentration appeared to be well correlated with total cholesterol, triglycerides, LDL-cholesterol, and apo B. These results were in contrast to total apo AIV, which was not different between dyslipidemic and normolipidemic individuals. The developed ELISA method for Lp B:AIVf in plasma combines specificity, reliability, and speed. The increase in Lp B:AIVf concentrations in various dyslipidemic states, together with a lack of change in total apo AIV concentrations, suggests a redistribution of apo AIV toward apo B-containing lipoproteins when these lipoproteins

  4. Genetic studies of human apolipoproteins. IX. Apolipoprotein D polymorphism and its relation to serum lipoprotein lipid levels.

    PubMed Central

    Kamboh, M I; Albers, J J; Majumder, P P; Ferrell, R E

    1989-01-01

    Apolipoprotein D (APO D) is a constituent of plasma high-density lipoproteins. Its precise role in lipid metabolism is not well established, though it may be involved in cholesterol esterification and cholester ester transport to the liver for catabolism. No genetic polymorphism has been reported in the APO D gene product. To investigate the extent of genetic variation at the APO D structural locus, we have developed an isoelectric focusing-immunoblotting technique and have screened a large number of plasma samples from U.S. whites, U.S. blacks, Nigerian blacks, the Aleuts of the Pribilof Islands, Eskimo groups from Kodiak Island and St. Lawrence Island, and Amerindian populations from Mexico and Canada. Except for the U.S. blacks and Nigerian blacks, the APO D locus is monomorphic in all other population groups tested. In populations with black ancestry, the products of two alleles, APO D*1 and APO D*2, have been observed at respective allele frequencies .987 and .013 in U.S. blacks and .978 and .022 in Nigerian blacks. The detection of a unique protein polymorphism in blacks makes APO D a useful black marker of significance in anthropogenetics and racial admixture studies. In addition to the interindividual variation observed, APO D reveals extensive intraindividual molecular variation with a multiple banding pattern. The basis of this molecular variation is explained, in part, by variation in the number of terminal sialic acid residues. We have investigated the effect of the APO D polymorphism on triglycerides, total cholesterol, LDL-, VLDL-, HDL-, and HDL3 cholesterol in 352 Nigerian blacks (190 males and 162 females).(ABSTRACT TRUNCATED AT 250 WORDS) Images Figure 1 Figure 2 PMID:2741945

  5. Human apolipoprotein B transgenic SHR/NDmcr-cp rats show exacerbated kidney dysfunction.

    PubMed

    Asahina, Makoto; Shimizu, Fumi; Ohta, Masayuki; Takeyama, Michiyasu; Tozawa, Ryuichi

    2015-01-01

    Nephropathy frequently co-occurs with metabolic syndrome in humans. Metabolic syndrome is a cluster of metabolic diseases including obesity, diabetes, hypertension, and dyslipidemia, and some previous studies revealed that dyslipidemia contributes to the progression of kidney dysfunction. To establish a new nephropathy model with metabolic syndrome, we produced human apolipoprotein B (apoB) transgenic (Tg.) SHR/NDmcr-cp (SHR-cp/cp) rats, in which dyslipidemia is exacerbated more than in an established metabolic syndrome model, SHR-cp/cp rats. Human apoB Tg. SHR-cp/cp rats showed obesity, hyperinsulinemia, hypertension, and severe hyperlipidemia. They also exhibited exacerbated early-onset proteinuria, accompanied by increased kidney injury and increased oxidative and inflammatory markers. Histological analyses revealed the characteristic features of human apoB Tg. SHR-cp/cp rats including prominent glomerulosclerosis with lipid accumulation. Our newly established human apoB Tg. SHR-cp/cp rat could be a useful model for the nephropathy in metabolic syndrome and for understanding the interaction between dyslipidemia and renal dysfunction in metabolic syndrome.

  6. Human apolipoprotein B transgenic SHR/NDmcr-cp rats show exacerbated kidney dysfunction

    PubMed Central

    ASAHINA, Makoto; SHIMIZU, Fumi; OHTA, Masayuki; TAKEYAMA, Michiyasu; TOZAWA, Ryuichi

    2015-01-01

    Nephropathy frequently co-occurs with metabolic syndrome in humans. Metabolic syndrome is a cluster of metabolic diseases including obesity, diabetes, hypertension, and dyslipidemia, and some previous studies revealed that dyslipidemia contributes to the progression of kidney dysfunction. To establish a new nephropathy model with metabolic syndrome, we produced human apolipoprotein B (apoB) transgenic (Tg.) SHR/NDmcr-cp (SHR-cp/cp) rats, in which dyslipidemia is exacerbated more than in an established metabolic syndrome model, SHR-cp/cp rats. Human apoB Tg. SHR-cp/cp rats showed obesity, hyperinsulinemia, hypertension, and severe hyperlipidemia. They also exhibited exacerbated early-onset proteinuria, accompanied by increased kidney injury and increased oxidative and inflammatory markers. Histological analyses revealed the characteristic features of human apoB Tg. SHR-cp/cp rats including prominent glomerulosclerosis with lipid accumulation. Our newly established human apoB Tg. SHR-cp/cp rat could be a useful model for the nephropathy in metabolic syndrome and for understanding the interaction between dyslipidemia and renal dysfunction in metabolic syndrome. PMID:25912321

  7. The apolipoprotein A-I mimetic peptide, D-4F, alleviates ox-LDL-induced oxidative stress and promotes endothelial repair through the eNOS/HO-1 pathway.

    PubMed

    Liu, Donghui; Ding, Zhenzhen; Wu, Mengzhang; Xu, Wenqi; Qian, Mingming; Du, Qian; Zhang, Le; Cui, Ye; Zheng, Jianlan; Chang, He; Huang, Caihua; Lin, Donghai; Wang, Yan

    2017-04-01

    Apolipoprotein A-I (apoA-I) mimetic peptide exerts many anti-atherogenic properties. However, the underlying mechanisms related to the endothelial protective effects remain elusive. In this study, the apoA-I mimetic peptide, D-4F, was used. Proliferation assay, wound healing, and transwell migration experiments showed that D-4F improved the impaired endothelial proliferation and migration resulting from ox-LDL. Endothelial adhesion molecules expression and monocyte adhesion assay demonstrated that D-4F inhibited endothelial inflammation. Caspase-3 activation and TUNEL stain indicated that D-4F reduced endothelial cell apoptosis. A pivotal anti-oxidant enzyme, heme oxygenase-1 (HO-1) was upregulated by D-4F. The Akt/AMPK/eNOS pathways were involved in the expression of HO-1 induced by D-4F. Moreover, the anti-oxidation, pro-proliferation, and pro-migration capacities of D-4F were diminished by the inhibitors of both eNOS (L-NAME) and HO-1 (Znpp). Additionally, downregulation of ATP-binding cassette transporter A1 (ABCA1) by siRNA abolished the activation of Akt, AMPK and eNOS, and reduced the upregulation of HO-1 triggered by D-4F. Furthermore, D-4F promoted the reendothelialization of injured intima in carotid artery injury model of C57BL/6J mice in vivo. In summary, these findings suggested that D-4F might be a powerful candidate in the protection of endothelial cells and the prevention of cardiovascular disease (CVD). Copyright © 2017. Published by Elsevier Ltd.

  8. Case report: A novel apolipoprotein A-I missense mutation apoA-I (Arg149Ser)Boston associated with decreased lecithin-cholesterol acyltransferase activation and cellular cholesterol efflux.

    PubMed

    Anthanont, Pimjai; Asztalos, Bela F; Polisecki, Eliana; Zachariah, Benoy; Schaefer, Ernst J

    2015-01-01

    We report a novel heterozygous apolipoprotein A-I (apoA-I) missense mutation (c.517C>A, p.Arg149Ser, designated as apoA-IBoston) in a 67-year-old woman and her 2 sons, who had mean serum high-density lipoprotein (HDL) cholesterol, apoA-I, and apoA-I in very large α-1 HDL that were 10%, 35%, and 16% of normal, respectively (all P < .05). The percentage of HDL cholesterol in the esterified form was also significantly (P < .05) reduced to 52% of control values. Cholesteryl ester tranfer protein (CETP) activity was normal. The mean global, adenosine triphosphate (ATP)-binding cassette transporter A1 and scavenger receptor B type I-mediated cellular cholesterol efflux capacity in apoB-depleted serum from affected family members were 41%, 37%, 47%, 54%, and 48% of control values, respectively (all P < .05). lecithin-cholesterol acyltransferase (LCAT) activity in plasma was 71% of controls, whereas in the cell-based assay, it was 73% of control values (P < .05). The data indicate that this novel apoA-I missense is associated with markedly decreased levels of HDL cholesterol and very large α-1 HDL, as well as decreased serum cellular cholesterol efflux and LCAT activity, but not with premature coronary heart disease, similar to other apoA-I mutations that have been associated with decreased LCAT activity. Copyright © 2015 National Lipid Association. Published by Elsevier Inc. All rights reserved.

  9. High-density lipoprotein and apolipoprotein A-I inhibit palmitate-induced translocation of toll-like receptor 4 into lipid rafts and inflammatory cytokines in 3T3-L1 adipocytes.

    PubMed

    Yamada, Hodaka; Umemoto, Tomio; Kawano, Mikihiko; Kawakami, Masanobu; Kakei, Masafumi; Momomura, Shin-Ichi; Ishikawa, San-E; Hara, Kazuo

    2017-03-04

    Saturated fatty acids (SFAs) activate toll-like receptor 4 (TLR4) signal transduction in macrophages and are involved in the chronic inflammation accompanying obesity. High-density lipoprotein (HDL) and apolipoprotein A-I (apoA-I) produce anti-inflammatory effects via reverse cholesterol transport. However, the underlying mechanisms by which HDL and apoA-I inhibit inflammatory responses in adipocytes remain to be determined. Here we examined whether palmitate increases the translocation of TLR4 into lipid rafts and whether HDL and apoA-I inhibit inflammation in adipocytes. Palmitate exposure (250 μM, 24 h) increased interleukin-6 and tumor necrosis factor-α gene expressions and translocation of TLR4 into lipid rafts in 3T3-L1 adipocytes. Pretreatment with HDL and apoA-I (50 μg/mL, 6 h) suppressed palmitate-induced inflammatory cytokine expression and TLR4 translocation into lipid rafts. Moreover, HDL and apoA-I inhibited palmitate-induced phosphorylation of nuclear factor-kappa B. HDL showed an anti-inflammatory effect via ATP-binding cassette transporter G1 and scavenger receptor class B, member 1, whereas apoA-I showed an effect via ATP-binding cassette transporter A1. These results demonstrated that HDL and apoA-I reduced palmitate-potentiated TLR4 trafficking into lipid rafts and its related inflammation in adipocytes via these specific transporters. Copyright © 2017. Published by Elsevier Inc.

  10. A Comparison of the Theoretical Relationship between HDL Size and the Ratio of HDL Cholesterol to Apolipoprotein A-I with Experimental Results from the Women’s Health Study

    PubMed Central

    Mazer, Norman A.; Giulianini, Franco; Paynter, Nina P.; Jordan, Paul; Mora, Samia

    2013-01-01

    Background HDL size and composition vary among individuals and may be associated with cardiovascular disease and diabetes. We investigated the theoretical relationship between HDL size and composition using an updated version of the spherical model of lipoprotein structure proposed by Shen et al. and compared its predictions with experimental data from the Women’s Health Study (WHS). Methods The Shen model was updated to predict the relationship between HDL diameter and the ratio of HDL-cholesterol (HDL-C) to apolipoprotein A-I (ApoA-I) plasma concentrations, i.e., the HDL-C/ApoA-I ratio. In WHS (n=26,772), NMR spectroscopy was used to measure the average HDL diameter (davg,NMR) and particle concentration (HDL-P); HDL-C and ApoA-I (mg/dL) were measured by standardized assays. Results The updated Shen model predicts a quasi-linear increase of HDL diameter with the HDL-C/ApoA-I ratio, consistent with the measured davg,NMR values from WHS, which ranged between 8.0 and 10.8 nm and correlated positively with the HDL-C/ApoA-I ratio (r=0.608, p<2.2×10−16). The WHS data were further described by a linear regression equation: dWHS (nm) = 4.66 + 12.31 HDL-C/ApoA-I. The validity of this equation for estimating HDL size was assessed with data from CETP deficiency and pharmacologic inhibition. We also illustrate how HDL-P can be estimated from the HDL size and ApoA-I level. Conclusions This study provides a large-scale experimental examination of the updated Shen model, offers new insights into HDL structure, composition and remodeling, and suggests that the HDL-C/ApoA-I ratio could be a readily available biomarker for estimating HDL size and HDL-P. PMID:23426429

  11. Role of apolipoprotein A-II in the structure and remodeling of human high-density lipoprotein (HDL): protein conformational ensemble on HDL.

    PubMed

    Gao, Xuan; Yuan, Shujun; Jayaraman, Shobini; Gursky, Olga

    2012-06-12

    High-density lipoproteins (HDL, or "good cholesterol") are heterogeneous nanoparticles that remove excess cell cholesterol and protect against atherosclerosis. The cardioprotective action of HDL and its major protein, apolipoprotein A-I (apoA-I), is well-established, yet the function of the second major protein, apolipoprotein A-II (apoA-II), is less clear. In this review, we postulate an ensemble of apolipoprotein conformations on various HDL. This ensemble is based on the crystal structure of Δ(185-243)apoA-I determined by Mei and Atkinson combined with the "double-hairpin" conformation of apoA-II(dimer) proposed in the cross-linking studies by Silva's team, and is supported by the wide array of low-resolution structural, biophysical, and biochemical data obtained by many teams over decades. The proposed conformational ensemble helps integrate and improve several existing HDL models, including the "buckle-belt" conformation of apoA-I on the midsize disks and the "trefoil/tetrafoil" arrangement on spherical HDL. This ensemble prompts us to hypothesize that endogenous apoA-II (i) helps confer lipid surface curvature during conversion of nascent discoidal HDL(A-I) and HDL(A-II) containing either apoA-I or apoA-II to mature spherical HDL(A-I/A-II) containing both proteins, and (ii) hinders remodeling of HDL(A-I/A-II) by hindering the expansion of the apoA-I conformation. Also, we report that, although endogenous apoA-II circulates mainly on the midsize spherical HDL(A-I/A-II), exogenous apoA-II can bind to HDL of any size, thereby slightly increasing this size and stabilizing the HDL assembly. This suggests distinctly different effects of the endogenous and exogenous apoA-II on HDL. Taken together, the existing results and models prompt us to postulate a new structural and functional role of apoA-II on human HDL.

  12. Topology of human apolipoprotein E3 uniquely regulates its diverse biological functions

    PubMed Central

    Chen, Jianglei; Li, Qianqian; Wang, Jianjun

    2011-01-01

    Human apolipoprotein E (apoE) is one of the major determinants in lipid transport, playing a critical role in atherosclerosis and other diseases. Binding to lipid and heparan sulfate proteoglycans (HSPG) induces apoE to adopt active conformations for binding to low-density lipoprotein receptor (LDLR) family. ApoE also interacts with beta amyloid peptide, manifests critical isoform-specific effects on Alzheimer’s disease. Despite the importance of apoE in these major human diseases, the fundamental questions of how apoE adjusts its structure upon binding to regulate its diverse functions remain unsolved. We report the NMR structure of apoE3, displaying a unique topology of three structural domains. The C-terminal domain presents a large exposed hydrophobic surface that likely initiates interactions with lipids, HSPG, and beta amyloid peptides. The unique topology precisely regulates apoE tertiary structure to permit only one possible conformational adaptation upon binding and provides a double security in preventing lipid-free and partially-lipidated apoE from premature binding to apoE receptors during receptor biogenesis. This topology further ensures the optimal receptor-binding activity by the fully lipidated apoE during lipoprotein transport in circulation and in the brain. These findings provide a structural framework for understanding the structural basis of the diverse functions of this important protein in human diseases. PMID:21873229

  13. Production of human apolipoprotein(a) transgenic NIBS miniature pigs by somatic cell nuclear transfer.

    PubMed

    Shimatsu, Yoshiki; Horii, Wataru; Nunoya, Tetsuo; Iwata, Akira; Fan, Jianglin; Ozawa, Masayuki

    2016-01-01

    Most cases of ischemic heart disease and stroke occur as a result of atherosclerosis. The purpose of this study was to produce a new Nippon Institute for Biological Science (NIBS) miniature pig model by somatic cell nuclear transfer (SCNT) for studying atherosclerosis. The human apolipoprotein(a) (apo(a)) genes were transfected into kidney epithelial cells derived from a male and a female piglet. Male cells were used as donors initially, and 275 embryos were transferred to surrogates. Three offspring were delivered, and the production efficiency was 1.1% (3/275). Serial female cells were injected into 937 enucleated oocytes. Eight offspring were delivered (production efficiency: 0.9%) from surrogates. One male and 2 female transgenic miniature pigs matured well. Lipoprotein(a) was found in the male and one of the female transgenic animals. These results demonstrate successful production of human apo(a) transgenic NIBS miniature pigs by SCNT. Our goal is to establish a human apo(a) transgenic NIBS miniature pig colony for studying atherosclerosis.

  14. No severe bottleneck during human evolution: evidence from two apolipoprotein C-II deficiency alleles.

    PubMed Central

    Xiong, W J; Li, W H; Posner, I; Yamamura, T; Yamamoto, A; Gotto, A M; Chan, L

    1991-01-01

    The DNA sequences of a Japanese and a Venezuelan apolipoprotein (apo) C-II deficiency allele, of a normal Japanese apo C-II gene, and of a chimpanzee apo C-II gene were amplified by PCR, and their nucleotide sequences were determined on multiple clones of the PCR products. The normal Japanese sequence is identical to--and the chimpanzee sequence differs by only three nucleotides from--a previously published normal Caucasian sequence. In contrast, the two human mutant sequences each differ from the normal apo C-II gene sequence by several nucleotides, including deletions. The data suggest that both mutant alleles arose greater than 500,000 years ago. It is shown that a defective allele can persist in a population for only a short time if a bottleneck occurs. Therefore, the antiquity of the two alleles suggests no severe bottleneck during human evolution. Moreover, the fact that one allele is from Japan and the other is from a Venezuelan Caucasian family is more consistent with the multiregional evolution model of modern human origins than with the complete replacement or "out of Africa" model. PMID:1990844

  15. Apolipoprotein A5: A newly identified gene impacting plasmatriglyceride levels in humans and mice

    SciTech Connect

    Pennacchio, Len A.; Rubin, Edward M.

    2002-09-15

    Apolipoprotein A5 (APOA5) is a newly described member of theapolipoprotein gene family whose initial discovery arose from comparativesequence analysis of the mammalian APOA1/C3/A4 gene cluster. Functionalstudies in mice indicated that alteration in the level of APOA5significantly impacted plasma triglyceride concentrations. Miceover-expressing human APOA5 displayed significantly reducedtriglycerides, while mice lacking apoA5 had a large increase in thislipid parameter. Studies in humans have also suggested an important rolefor APOA5 in determining plasma triglyceride concentrations. In theseexperiments, polymorphisms in the human gene were found to define severalcommon haplotypes that were associated with significant changes intriglyceride concentrations in multiple populations. Several separateclinical studies have provided consistent and strong support for theeffect with 24 percent of Caucasians, 35 percent of African-Americans and53 percent of Hispanics carrying APOA5 haplotypes associated withincreased plasma triglyceride levels. In summary, APOA5 represents anewly discovered gene involved in triglyceride metabolism in both humansand mice whose mechanism of action remains to be deciphered.

  16. Production of Cloned Miniature Pigs Expressing High Levels of Human Apolipoprotein(a) in Plasma

    PubMed Central

    Ozawa, Masayuki; Himaki, Takehiro; Ookutsu, Shoji; Mizobe, Yamato; Ogawa, Junki; Miyoshi, Kazuchika; Yabuki, Akira; Fan, Jianglin; Yoshida, Mitsutoshi

    2015-01-01

    High lipoprotein(a) [Lp(a)] levels are a major risk factor for the development of atherosclerosis. However, because apolipoprotein(a) [apo(a)], the unique component of Lp(a), is found only in primates and humans, the study of human Lp(a) has been hampered due to the lack of appropriate animal models. Using somatic cell nuclear transfer (SCNT) techniques, we produced transgenic miniature pigs expressing human apo(a) in the plasma. First, we placed the hemagglutinin (HA)-tagged cDNA of human apo(a) under the control of the β-actin promoter and cytomegalovirus enhancer, and then introduced this construct into kidney epithelial cells. Immunostaining of cells with anti-HA antibody allowed identification of cells stably expressing apo(a); one of the positive clones was used to provide donor cells for SCNT, yielding blastocysts that expressed apo(a). Immunohistochemical analysis of tissue sections and RT-PCR analysis of total RNA from organs of cloned piglet revealed that apo(a) is expressed in various tissues/organs including heart, liver, kidney, and intestine. More importantly, a transgenic line exhibited a high level (>400 mg/dL) of Lp(a) in plasma, and the transgenic apo(a) gene was transmitted to the offspring. Thus, we generated a human apo(a)–transgenic miniature pig that can be used as a model system to study advanced atherosclerosis related to human disease. The anatomical and physiological similarities between the swine and human cardiovascular systems will make this pig model a valuable source of information on the role of apo(a) in the formation of atherosclerosis, as well as the mechanisms underlying vascular health and disease. PMID:26147378

  17. Internet AIS

    NASA Astrophysics Data System (ADS)

    Filjar, Renato; Desic, Sasa; Pokrajac, Danijela; Cubic, Ivica

    2005-05-01

    Automatic Identification System (AIS) has recently become the leading issue in maritime navigation and traffic management worldwide. The present AIS solution, based on a VHF data communications scheme, provides AIS functionalities for SOLAS (AIS Class A) vessels only in a limited environment defined by radio propagation properties. Here we present a novel approach in AIS development based on current mobile communication technologies. It utilises existing mobile communications equipment that the majority of targetted end-users own and are familiar with. A novel AIS concept aims to offer a transition of AIS data traffic to mobile Internet. An innovative AIS architecture supports AIS data processing, storing and transferring to authorised parties. This enhances not only the operational area, but also provides the global AIS with data transfer security and an improved aids-for-navigation service, with all legally traceable vessels (both AIS Class A and AIS Class B) included in the system. In order to provide the development framework for Internet AIS, a set of essential four use-cases, a communication protocol and the first Internet AIS prototype have been recently developed and are briefly introduced in this article.

  18. Macrophage-specific expression of human apolipoprotein E reduces atherosclerosis in hypercholesterolemic apolipoprotein E-null mice.

    PubMed Central

    Bellosta, S; Mahley, R W; Sanan, D A; Murata, J; Newland, D L; Taylor, J M; Pitas, R E

    1995-01-01

    apoE deficiency causes hyperlipidemia and premature atherosclerosis. To determine if macrophage-specific expression of apoE would decrease the extent of atherosclerosis, we expressed human apoE in macrophages of apoE-null mice (apoE-/-) and assessed the effect on lipid accumulation in cells of the arterial wall. Macrophage-specific expression of human apoE in normal mice was obtained by use of the visna virus LTR. These animals were bred with apoE-/- mice to produce animals hemizygous for expression of human apoE in macrophages in the absence of murine apoE (apoE-/-,hTgE+/0). Low levels of human apoE mRNA were present in liver and spleen and high levels in lung and peritoneal macrophages. Human apoE was secreted by peritoneal macrophages and was detected in Kupffer cells of the liver. Human apoE in the plasma of apoE-/-,hTgE+/0 mice (n = 30) was inversely correlated (P < 0.005) with the plasma cholesterol concentration. After 15 wk on a normal chow diet, atherosclerosis was assessed in apoE-/-,hTgE+/0 animals and in apoE-/-,hTgE0/0 littermates matched for plasma cholesterol level (approximately 450 mg/dl) and lipoprotein profile. There was significantly less atherosclerosis in both the aortic sinus and in the proximal aorta (P < 0.0001) in the animals expressing the human apoE transgene. In apo-E-/-,hTgE+/0 animals, which had detectable atherosclerotic lesions, human apoE was detected in the secretory apparatus of macrophage-derived foam cells in the arterial wall. The data demonstrate that expression of apoE by macrophages is antiatherogenic even in the presence of high levels of atherogenic lipoproteins. The data suggest that apoE prevents atherosclerosis by promoting cholesterol efflux from cells of the arterial wall. Images PMID:7593602

  19. Niacin increases HDL biogenesis by enhancing DR4-dependent transcription of ABCA1 and lipidation of apolipoprotein A-I in HepG2 cells

    PubMed Central

    Zhang, Lin-Hua; Kamanna, Vaijinath S.; Ganji, Shobha H.; Xiong, Xi-Ming; Kashyap, Moti L.

    2012-01-01

    The lipidation of apoA-I in liver greatly influences HDL biogenesis and plasma HDL levels by stabilizing the secreted apoA-I. Niacin is the most effective lipid-regulating agent clinically available to raise HDL. This study was undertaken to identify regulatory mechanisms of niacin action in hepatic lipidation of apoA-I, a critical event involved in HDL biogenesis. In cultured human hepatocytes (HepG2), niacin increased: association of apoA-I with phospholipids and cholesterol by 46% and 23% respectively, formation of lipid-poor single apoA-I molecule-containing particles up to ∼ 2.4-fold, and pre β 1 and α migrating HDL particles. Niacin dose-dependently stimulated the cell efflux of phospholipid and cholesterol and increased transcription of ABCA1 gene and ABCA1 protein. Mutated DR4, a binding site for nuclear factor liver X receptor alpha (LXR α ) in the ABCA1 promoter, abolished niacin stimulatory effect. Further, knocking down LXR α or ABCA1 by RNA interference eliminated niacin-stimulated apoA-I lipidation. Niacin treatment did not change apoA-I gene expression. The present data indicate that niacin increases apoA-I lipidation by enhancing lipid efflux through a DR4-dependent transcription of ABCA1 gene in HepG2 cells. A stimulatory role of niacin in early hepatic formation of HDL particles suggests a new mechanism that contributes to niacin action to increase the stability of newly synthesized circulating HDL. PMID:22389325

  20. Dietary fat elevates hepatic apoA-I production by increasing the fraction of apolipoprotein A-I mRNA in the translating pool.

    PubMed

    Azrolan, N; Odaka, H; Breslow, J L; Fisher, E A

    1995-08-25

    Elevated plasma high density lipoprotein cholesterol (HDL-C) levels are associated with a decreased risk for coronary heart disease. Ironically, diets enriched in saturated fat and cholesterol (HF/HC diets), which tend to accelerate atherosclerotic processes by increasing LDL cholesterol levels, also raise HDL-C. We have recently reported, using a human apoA-I (hapoA-1) transgenic mouse model, that the elevation of HDL-C by a HF/HC diet is attributable, in part, to an increase in the hepatic production of hapoA-1. To further define the hepatocellular processes associated with this induction, we have prepared primary hepatocytes from hapoA-1 transgenic mice. Rates of hapoA-1 secretion were 40% greater from cells prepared from animals fed the HF/HC relative to a low fat-low cholesterol (LF/LC) control diet. The abundance of hapoA-1 mRNA in these cells was similar between hepatocytes prepared from the HF/HC and LF/LC diet fed animals, suggesting a post-transcriptional mechanism that does not involve mRNA stability. Inhibition of secretion using brefeldin A revealed an increase in cellular hapoA-1 accumulation. Thus, the HF/HC diet apparently affects hepatic hapoA-1 production via a mechanism that is manifest prior to the exit of newly synthesized hapoA-1 from the Golgi. Pulse-chase experiments revealed a 39% greater peak hapoA-1 synthesis, with no difference in the degradation of total labeled hapoA-1 protein, as a result of the HF/HC diet feeding. Finally, resolution of liver S10 extracts via sucrose density sedimentation and metrizamide density equilibrium gradient centrifugation analyses both revealed similar increases (31 and 24%, respectively) in the relative percentage of hapoA-1 mRNA associated with the translating polysomal fractions as a result of the HF/HC feeding. Together, these data suggest that the HF/HC diet affects hepatic hapoA-1 production via a specific modulation in the relative amount of hapoA-1 mRNA in the polysomal pool. These observations

  1. Human-centered automation and AI - Ideas, insights, and issues from the Intelligent Cockpit Aids research effort

    NASA Technical Reports Server (NTRS)

    Abbott, Kathy H.; Schutte, Paul C.

    1989-01-01

    A development status evaluation is presented for the NASA-Langley Intelligent Cockpit Aids research program, which encompasses AI, human/machine interfaces, and conventional automation. Attention is being given to decision-aiding concepts for human-centered automation, with emphasis on inflight subsystem fault management, inflight mission replanning, and communications management. The cockpit envisioned is for advanced commercial transport aircraft.

  2. Distinctive structure and interfacial activity of the human apolipoprotein A-IV 347S isoprotein

    PubMed Central

    Weinberg, Richard B.; Cook, Victoria R.

    2010-01-01

    The T347S polymorphism in the human apolipoprotein (apo) A-IV gene is present at high frequencies among all the world's populations. Carriers of a 347S allele exhibit faster clearance of triglyceride-rich lipoproteins, greater adiposity, and increased risk for developing atherosclerosis, which suggests that this conservative amino acid substitution alters the structure of apo A-IV. Herein we have used spectroscopic and surface chemistry techniques to examine the structure, stability, and interfacial properties of the apo A-IV 347S isoprotein. Circular dichroism spectroscopy revealed that the 347S isoprotein has similar α-helical structure but lower thermodynamic stability than the 347T isoprotein. Fluorescence spectroscopy found that the 347S isoprotein exhibits an enhanced tyrosine emission and reduced tyrosine→tryptophan energy transfer, and second derivative UV absorption spectra noted increased tyrosine exposure, suggesting that the 347S isoprotein adopts a looser tertiary conformation. Surface chemistry studies found that although the 347S isoprotein bound rapidly to the lipid interface, it has a lower interfacial exclusion pressure and lower elastic modulus than the 347T isoprotein. Together, these observations establish that the T347S substitution alters the conformation of apo A-IV and lowers its interfacial activity—changes that could account for the effect of this polymorphism on postprandial lipid metabolism. PMID:20554794

  3. Expression of the human apolipoprotein E gene suppresses steroidogenesis in mouse Y1 adrenal cells

    SciTech Connect

    Reyland, M.E.; Forgez, P.; Prack, M.M.; Williams, D.L. ); Gwynne, J.T. )

    1991-03-15

    The lipid transport protein, apolipoprotein E (apoE), is expressed in many peripheral tissues in vivo including the adrenal gland and testes. To investigate the role of apoE in adrenal cholesterol homeostasis, the authors have expressed a human apoE genomic clone in the Y1 mouse adrenocortical cell line. Y1 cells do not express endogenous apoE mRNA or protein. Expression of apoE in Y1 cells resulted in a dramatic decrease in basal steroidogenesis; secretion of fluorogenic steroid was reduced 7- to {gt}100-fold relative to Y1 parent cells. Addition of 5-cholesten-3{beta},25-idol failed to overcome the suppression of steroidogenesis in these cells. Cholesterol esterification under basal conditions, as measured by the production of cholesteryl ({sup 14}C)oleate, was similar in the Y1 parent and the apoE-transfected cell lines. Upon incubation with adrenocorticotropin or dibutyryl cAMP, production of cholesteryl ({sup 14}C)oleate decreased 5-fold in the Y1 parent cells but was unchanged in the apoE-transfected cell lines. These results suggest that apoE may be an important modulator of cholesterol utilization and steroidogenesis in adrenal cells.

  4. Production of the kringle fragments of human apolipoprotein(a) by continuous lactose induction strategy.

    PubMed

    Lim, Hyung-Kwon; Kim, Sung-Geun; Jung, Kyung-Hwan; Seo, Jin-Ho

    2004-03-18

    A novel lactose induction strategy for the production of rhLK68, the kringle fragments of human apolipoprotein(a) (apo(a)) as a novel anti-angiogenic protein, was investigated. A scale-up of the production was accompanied by a decrease in expression level, and severe aggregation occurred during the solubilization of rhLK68 from the inclusion body during a conventional single introduction of lactose. To overcome this problem, a continuous induction strategy was applied where lactose was mixed with glycerol and fed continuously in a dissolved oxygen (DO)-stat manner. With the sub-optimal feed medium consisted of 1:50 of lactose/glycerol (w/w), the expression level reached 16% of the total cellular protein, which was 1.6-fold higher than that obtained from the conventional lactose induction. Moreover, the solubilization yield of rhLK68 from the inclusion body increased from 30 +/- 5 to 85 +/- 3% compared to the conventional single introduction of lactose. This result suggests that the continuous lactose induction strategy beneficially influenced the expression level of rhLK68 and the quality of its inclusion body.

  5. Comparative surface antimicrobial properties of synthetic biocides and novel human apolipoprotein E derived antimicrobial peptides.

    PubMed

    Forbes, Sarah; McBain, Andrew J; Felton-Smith, Susan; Jowitt, Thomas A; Birchenough, Holly L; Dobson, Curtis B

    2013-07-01

    Medical device infection remains a major clinical concern. Biocidal compounds have been incorporated into medical device materials ideally to inhibit bacterial colonisation whilst exhibiting relatively low cytotoxicity. We compared the antibacterial activity, anti-biofilm efficacy and cytotoxicity of a novel peptide derivative of human apolipoprotein E (apoEdpL-W) to that of commonly used biocides, before and after coating onto a range of standard polymers. Since the antimicrobial function of most biocides frequently involves associations with cellular membranes, we have also studied the detailed interactions of the test antimicrobials with phospholipid bilayers, using the quartz crystal microbalance device combined with dual-polarisation interferometry. ApoEdpL-W displayed broad-spectrum antibacterial activity and marked efficacy against nascent Staphylococcus aureus biofilms. Compounds showed better antimicrobial activity when combined with hydrogel materials than with non-porous materials. The membrane interactions of apoEdpL-W were most similar to that of PHMB, with both agents appearing to readily bind and insert into lipid bilayers, possibly forming pores. However apoEdpL-W showed lower cytotoxicity than PHMB, its efficacy was less affected by the presence of serum, and it demonstrated the highest level of biocompatibility of all the biocides, as indicated by our measurement of its antimicrobial biocompatibility index. This work shows the potential of apoEdpL-W as an effective antiseptic coating agent.

  6. The insertion of human apolipoprotein H into phospholipid membranes: a monolayer study.

    PubMed

    Wang, S X; Cai, G P; Sui, S F

    1998-10-15

    Apolipoprotein H (ApoH) is a plasma glycoprotein isolated from human serum. The interactions of ApoH with lipid membrane were reported to be essential for its physiological and pathogenic roles. In this paper we studied the ability of ApoH to insert into phospholipid membranes using the monolayer approach. The results show that ApoH is surface active and can insert into the lipid monolayers. The insertion ability of ApoH is stronger when a higher content of negatively charged lipids is present in the membrane. The acidic-pH and low-ionic-strength conditions will also enhance ApoH insertion, but these factors may not have much influence on the final insertion ability of ApoH, suggesting that, in the mechanism of ApoH insertion, not only electrostatic forces, but also hydrophobic interactions, are evidently involved. Modification by heat inactivation and reduction/alkylation does not change the critical insertion pressure (pic) of ApoH, suggesting a stable domain, maybe a linear sequence motif, but not the native three-dimensional structure of ApoH, is responsible for its insertion. The extent to which insertion of ApoH into phospholipid membranes may facilitate the 'immune cleaning' of plasma liposomes is discussed.

  7. Isolation of a cDNA clone encoding the amino-terminal region of human apolipoprotein B

    SciTech Connect

    Protter, A.A.; Hardman, D.A.; Schilling, J.W.; Miller, J.; Appleby, V.; Chen, G.C.; Kirsher, S.W.; McEnroe, G.; Kane, J.P.

    1986-03-01

    A partial cDNA clone for the B-26 region of apolipoprotein B was isolated from an adult human liver DNA library by screening with an oligonucleotide probe derived from amino-terminal protein sequence obtained from purified B-26 peptide. Antisera against a synthetic 17-residue peptide whose amino acid sequence was encoded by the clone cross-reacts with apolipoproteins B-26, B-100, and B-48, but not with B-74. The nucleotide sequence immediately upstream from the amino terminus of B-26 codes for an apparent signal sequence, implying that the B-26 moiety is in an amino-terminal locus in the B-100 protein. That this sequence represents a 5' end region is further supported by primer extension analysis using a fragment of the cDNA clone and by S1 nuclease protection experiments using the corresponding region in a genomic clone.

  8. Two independent apolipoprotein a5 Haplotypes influence human plasma triglyceride levels

    SciTech Connect

    Pennacchio, Len A.; Olivier, Michael; Hubacek, Jaroslav A.; Krauss, Ronald M.; Rubin, Edward M.; Cohen, Jonathan C.

    2002-09-16

    The recently identified apolipoprotein A5 gene (APOA5) has been shown to play an important role in determining plasma triglyceride concentrations in humans and mice. We previously identified an APOA5 haplotype (designated APOA5*2) that is present in {approx}16 percent of Caucasians and is associated with increased plasma triglyceride concentrations. In this report we describe another APOA5 haplotype (APOA5*3) containing the rare allele of the single nucleotide polymorphism c.56C>G that changes serine to tryptophan at codon 19 and is independently associated with high plasma triglyceride levels in three different populations. In a sample of 264 Caucasian men and women with plasma triglyceride concentrations above the 90th percentile or below the 10th percentile, the APOA5*3 haplotype was more than three-fold more common in the group with high plasma triglyceride levels. In a second independently ascertained sample of Caucasian men and women (n 1/4 419) who were studied while consuming their self-selected diets as well as after high-carbohydrate diets and high-fat diets, the APOA5*3 haplotype was associated with increased plasma triglyceride levels on all three dietary regimens. In a third population comprising 2660 randomly selected individuals, the APOA5*3 haplotype was found in 12 percent of Caucasians, 14 percent of African-Americans and 28 percent of Hispanics and was associated with increased plasma triglyceride levels in both men and women in each ethnic group. These findings establish that the APOA5 locus contributes significantly to inter-individual variation in plasma triglyceride levels in humans. Together, the APOA5*2 and APOA5*3 haplotypes are found in 25 50 percent of African-Americans, Hispanics and Caucasians and support the contribution of common human variation to quantitative phenotypes in the general population.

  9. Apolipoprotein A-I assayed in human serum by isotope dilution as a potential standard for immunoassay

    SciTech Connect

    Weech, P.K.; Jewer, D.; Marcel, Y.L.

    1988-01-01

    We measured the amount of apoA-I in serum by isotope dilution, finding 1.33 mg/ml (standard deviation 0.177) in six normolipidemic, healthy subjects. We developed this method by adapting published techniques to purify apoA-I from 3 ml of serum in two steps: density gradient ultracentrifugation and high performance liquid chromatography gel filtration. The 125I-labeled apoA-I tracer was first screened, by incubation with serum, to select labeled apoA-I which retained the ability to exchange with native apoA-I and bind to HDL. A known amount of 125I-labeled apoA-I-labeled HDL was added to unknown serum samples; apoA-I was reisolated from the serum and its specific radioactivity was used to calculate the dilution of the added, labeled apoA-I by the unlabeled apoA-I in the unknown serum. By not relying on immunochemical techniques, the isotope dilution assay provided results that are independent of the expression of individual apoA-I antigenic sites. Therefore, sera that have been assayed by isotope dilution can serve as standards to evaluate the accuracy of immunoassays for serum apoA-I and provide primary standards for such immunoassays.

  10. Structural basis for distinct functions of the naturally occurring Cys mutants of human apolipoprotein A-I[S

    PubMed Central

    Gursky, Olga; Jones, Martin K.; Mei, Xiaohu; Segrest, Jere P.; Atkinson, David

    2013-01-01

    HDL removes cell cholesterol and protects against atherosclerosis. ApoA-I provides a flexible structural scaffold and an important functional ligand on the HDL surface. We propose structural models for apoA-IMilano (R173C) and apoA-IParis (R151C) mutants that show high cardioprotection despite low HDL levels. Previous studies established that two apoA-I molecules encircle HDL in an antiparallel, helical double-belt conformation. Recently, we solved the atomic structure of lipid-free Δ(185–243)apoA-I and proposed a conformational ensemble for apoA-IWT on HDL. Here we modify this ensemble to understand how intermolecular disulfides involving C173 or C151 influence protein conformation. The double-belt conformations are modified by belt rotation, main-chain unhinging around Gly, and Pro-induced helical bending, and they are verified by comparison with previous experimental studies and by molecular dynamics simulations of apoA-IMilano homodimer. In our models, the molecular termini repack on various-sized HDL, while packing around helix-5 in apoA-IWT, helix-6 in apoA-IParis, or helix-7 in apoA-IMilano homodimer is largely conserved. We propose how the disulfide-induced constraints alter the protein conformation and facilitate dissociation of the C-terminal segment from HDL to recruit additional lipid. Our models unify previous studies of apoA-IMilano and demonstrate how the mutational effects propagate to the molecular termini, altering their conformations, dynamics, and function. PMID:24038317

  11. Complex effects of inhibiting hepatic apolipoprotein B100 synthesis in humans

    PubMed Central

    Reyes-Soffer, Gissette; Moon, Byoung; Hernandez-Ono, Antonio; Dionizovik-Dimanovski, Marija; Jimenez, Jhonsua; Obunike, Joseph; Thomas, Tiffany; Ngai, Colleen; Fontanez, Nelson; Donovan, Daniel S.; Karmally, Wahida; Holleran, Stephen; Ramakrishnan, Rajasekhar; Mittleman, Robert S.; Ginsberg, Henry N.

    2016-01-01

    Mipomersen is a 20mer antisense oligonucleotide (ASO) that inhibits apolipoprotein B (apoB) synthesis; its low-density lipoprotein (LDL)–lowering effects should therefore result from reduced secretion of very-low-density lipoprotein (VLDL). We enrolled 17 healthy volunteers who received placebo injections weekly for 3 weeks followed by mipomersen weekly for 7 to 9 weeks. Stable isotopes were used after each treatment to determine fractional catabolic rates and production rates of apoB in VLDL, IDL (intermediate-density lipoprotein), and LDL, and of triglycerides in VLDL. Mipomersen significantly reduced apoB in VLDL, IDL, and LDL, which was associated with increases in fractional catabolic rates of VLDL and LDL apoB and reductions in production rates of IDL and LDL apoB. Unexpectedly, the production rates of VLDL apoB and VLDL triglycerides were unaffected. Small interfering RNA–mediated knockdown of apoB expression in human liver cells demonstrated preservation of apoB secretion across a range of apoB synthesis. Titrated ASO knockdown of apoB mRNA in chow-fed mice preserved both apoB and triglyceride secretion. In contrast, titrated ASO knockdown of apoB mRNA in high-fat–fed mice resulted in stepwise reductions in both apoB and triglyceride secretion. Mipomersen lowered all apoB lipoproteins without reducing the production rate of either VLDL apoB or triglyceride. Our human data are consistent with longstanding models of posttranscriptional and posttranslational regulation of apoB secretion and are supported by in vitro and in vivo experiments. Targeting apoB synthesis may lower levels of apoB lipoproteins without necessarily reducing VLDL secretion, thereby lowering the risk of steatosis associated with this therapeutic strategy. PMID:26819195

  12. AI techniques for optimizing multi-objective reservoir operation upon human and riverine ecosystem demands

    NASA Astrophysics Data System (ADS)

    Tsai, Wen-Ping; Chang, Fi-John; Chang, Li-Chiu; Herricks, Edwin E.

    2015-11-01

    Flow regime is the key driver of the riverine ecology. This study proposes a novel hybrid methodology based on artificial intelligence (AI) techniques for quantifying riverine ecosystems requirements and delivering suitable flow regimes that sustain river and floodplain ecology through optimizing reservoir operation. This approach addresses issues to better fit riverine ecosystem requirements with existing human demands. We first explored and characterized the relationship between flow regimes and fish communities through a hybrid artificial neural network (ANN). Then the non-dominated sorting genetic algorithm II (NSGA-II) was established for river flow management over the Shihmen Reservoir in northern Taiwan. The ecosystem requirement took the form of maximizing fish diversity, which could be estimated by the hybrid ANN. The human requirement was to provide a higher satisfaction degree of water supply. The results demonstrated that the proposed methodology could offer a number of diversified alternative strategies for reservoir operation and improve reservoir operational strategies producing downstream flows that could meet both human and ecosystem needs. Applications that make this methodology attractive to water resources managers benefit from the wide spread of Pareto-front (optimal) solutions allowing decision makers to easily determine the best compromise through the trade-off between reservoir operational strategies for human and ecosystem needs.

  13. Effects of apolipoproteins on the kinetics of cholesterol exchange

    SciTech Connect

    Letizia, J.Y.; Phillips, M.C. )

    1991-01-22

    The effects of apolipoproteins on the kinetics of cholesterol exchange have been investigated by monitoring the transfer of ({sup 14}C)cholesterol from donor phospholipid/cholesterol complexes containing human apolipoproteins A, B, or C. Negatively charged discoidal and vesicular particles containing purified apolipoproteins complexed with lipid and a trace of ({sup 14}C)cholesterol were incubated with a 10-fold excess of neutral, acceptor, small unilamellar vesicles. The donor and acceptor particles were separated by chromatogrphy of DEAE-Sepharose, and the rate of movement of labeled cholesterol was analyzed as a first-order exchange process. The kinetics of exchange of cholesterol from both vesicular and discoidal complexes that contain apoproteins are consistent with an aqueous diffusion mechanism, as has been established previously for PC/cholesterol SUV. Apolipoproteins A-I, A-II, reduced and carboxymethylated A-11, and B-100 present in SUV at the same lipid/protein (w/w) ratio all enhance the rate of cholesterol exchange to about the same degree. Cholesterol molecules exchange more rapidly from discoidal complexes. Generally, as the diameter of apoprotein/phospholipid/cholesterol discs decreases, t{sub 1/2} for cholesterol exchange decreases. Since small bilayer discs have a relatively high ratio of boundary to face surface area, cholesterol molecules desorb more rapidly than from larger discs. The modulation of lipid packing by the apoprotein molecules present at the surface of lipoprotein particles affects the rate of cholesterol exchange from such particles.

  14. In Vivo Human Apolipoprotein E Isoform Fractional Turnover Rates in the CNS

    PubMed Central

    Patterson, Bruce W.; Pyatkivskyy, Yuriy; Kim, Jungsu; Yarasheski, Kevin E.; Wang, Jennifer X.; Mawuenyega, Kwasi G.; Jiang, Hong; Parsadanian, Maia; Yoon, Hyejin; Kasten, Tom; Sigurdson, Wendy C.; Xiong, Chengjie; Goate, Alison; Holtzman, David M.; Bateman, Randall J.

    2012-01-01

    Apolipoprotein E (ApoE) is the strongest genetic risk factor for Alzheimer’s disease and has been implicated in the risk for other neurological disorders. The three common ApoE isoforms (ApoE2, E3, and E4) each differ by a single amino acid, with ApoE4 increasing and ApoE2 decreasing the risk of Alzheimer’s disease (AD). Both the isoform and amount of ApoE in the brain modulate AD pathology by altering the extent of amyloid beta (Aβ) peptide deposition. Therefore, quantifying ApoE isoform production and clearance rates may advance our understanding of the role of ApoE in health and disease. To measure the kinetics of ApoE in the central nervous system (CNS), we applied in vivo stable isotope labeling to quantify the fractional turnover rates of ApoE isoforms in 18 cognitively-normal adults and in ApoE3 and ApoE4 targeted-replacement mice. No isoform-specific differences in CNS ApoE3 and ApoE4 turnover rates were observed when measured in human CSF or mouse brain. However, CNS and peripheral ApoE isoform turnover rates differed substantially, which is consistent with previous reports and suggests that the pathways responsible for ApoE metabolism are different in the CNS and the periphery. We also demonstrate a slower turnover rate for CSF ApoE than that for amyloid beta, another molecule critically important in AD pathogenesis. PMID:22675504

  15. Metabolism of Apolipoprotein A-II Containing Triglyceride Rich ApoB Lipoproteins in Humans

    PubMed Central

    Desai, Nirav K.; Ooi, Esther M.; Mitchell, Paul D.; Furtado, Jeremy; Sacks, Frank M.

    2015-01-01

    Objective To characterize human triglyceride-rich lipoproteins (TRL) with and without apoA-II and to study their metabolism in vivo. Methods Plasma from 11 participants on a controlled diet given a bolus infusion of [D5]L-phenylalanine to label apoB was combined into four pools and applied to anti-apoA-II immunoaffinity columns. Fractions with and without apoA-II were separated into VLDL and IDL by ultracentrifugation; lipids and apolipoproteins were measured. For kinetic measurements, apoB was isolated and hydrolyzed to the constituent amino acids. Tracer enrichment was measured by GCMS. Metabolic rates were determined by SAAM-II. Results VLDL and IDL with apoA-II comprised 7% and 9% of total VLDL and IDL apoB respectively. VLDL with apoA-II was enriched in apoC-III, apoE, and cholesterol compared to VLDL without apoA-II. Mean apoB FCR of VLDL with apoA-II was significantly lower than for VLDL without apoA-II (2.80±0.96 pools/day v.s. 5.09±1.69 pools/day, P=0.009). A higher percentage of VLDL with apoA-II was converted to IDL than was cleared from circulation, compared to VLDL without apoA-II (96±8% vs. 45±22%; P=0.007). The rate constants for conversion of VLDL to IDL were similar for VLDL with and without apoA-II. Thus, a very low rate constant for clearance accounted for the lower FCR of VLDL with apoA-II. Conclusion VLDL with apoA-II represents a small pool of VLDL particles that has a slow FCR and is predominantly converted to IDL rather than cleared from the circulation. PMID:26071654

  16. Separation of apolipoproteins of human very low density lipoproteins by chromatofocusing.

    PubMed

    März, W; Gross, W

    1983-07-01

    Chromatofocusing represents a new chromatographic procedure for the separation of proteins according to their isoelectric points. We describe the application of this method for the fractionation of the urea-soluble apolipoproteins of very low density lipoproteins. They were separated into five peaks, four of which were homogeneous as judged by polyacrylamide gel electrophoresis in the presence of 7 mol/l urea.

  17. Heredity links natural hazards and human health: Apolipoprotein E gene moderates the health of earthquake survivors.

    PubMed

    Daly, Michael; MacLachlan, Malcolm

    2011-03-01

    This study aimed to investigate the role of the apolipoprotein ε4 allele in moderating the influence of an exogenous stressor, an earthquake, on health. A "natural experiment" design was used where the interaction between the presence of the apolipoprotein ε4 allele and the level of subjective and objective exposure to a devastating earthquake was examined in a population-based cohort of elderly Taiwanese (N = 718). The cognitive-affective dimension of health was assessed by measures of perceived control and depression and functional limitations were assessed using measures of instrumental activities of daily living and mobility. Overall health status was gauged using a single-item measure of self-rated health. Those who experienced damage to their property or were forced to move from their homes (high objective exposure) demonstrated low levels of self-rated health and somewhat lower perceived control a year later, only if they were apolipoprotein ε4 carriers. Similarly, those who found the earthquake severely distressing (high subjective exposure) were shown to have low levels of functioning and low self-rated health a year later, only if they possessed the ε4 allele. Our findings suggest that genetic variation in the apolipoprotein E gene may modify the health effects of the exogenous stress of natural disaster exposure.

  18. Plasma clearance of human low-density lipoprotein in human apolipoprotein B transgenic mice is related to particle diameter.

    PubMed

    Berneis, Kaspar; Shames, David M; Blanche, Patricia J; La Belle, Michael; Rizzo, Manfredi; Krauss, Ronald M

    2004-04-01

    To test for intrinsic differences in metabolic properties of low-density lipoprotein (LDL) as a function of particle size, we examined the kinetic behavior of 6 human LDL fractions ranging in size from 251 to 265 A injected intravenously into human apolipoprotein (apo) B transgenic mice. A multicompartmental model was formulated and fitted to the data by standard nonlinear regression using the Simulation, Analysis and Modeling (SAAM II) program. Smaller sized LDL particles (251 to 257 A) demonstrated a significantly slower fractional catabolic rate (FCR) (0.050 +/- 0.045 h(-1)) compared with particles of larger size (262 to 265 A) (0.134 +/- -0.015 h(-1), P <.03), and there was a significant correlation between FCR and the peak LDL diameter of the injected fractions (R(2) =.71, P <.034). The sum of the equilibration parameters, k(2,1) and k(1,2), for smaller LDL (0.255 h(-1) and 0.105 h(-1), respectively) was significantly smaller than that for larger LDL (0.277 h(-1) and 0.248 h(-1), respectively; P <.01), indicative of slower intravascular-extravascular exchange for smaller LDL. Therefore in this mouse model, smaller LDL particles are cleared more slowly from plasma than larger LDL and are exchanged more slowly with the extravascular space. This might be due to compositional or structural features of smaller LDL that lead to retarded clearance.

  19. Sequence Diversity and Large-Scale Typing of SNPs in the Human Apolipoprotein E Gene

    PubMed Central

    Nickerson, Deborah A.; Taylor, Scott L.; Fullerton, Stephanie M.; Weiss, Kenneth M.; Clark, Andrew G.; Stengård, Jari H.; Salomaa, Veikko; Boerwinkle, Eric; Sing, Charles F.

    2000-01-01

    A common strategy for genotyping large samples begins with the characterization of human single nucleotide polymorphisms (SNPs) by sequencing candidate regions in a small sample for SNP discovery. This is usually followed by typing in a large sample those sites observed to vary in a smaller sample. We present results from a systematic investigation of variation at the human apolipoprotein E locus (APOE), as well as the evaluation of the two-tiered sampling strategy based on these data. We sequenced 5.5 kb spanning the entire APOE genomic region in a core sample of 72 individuals, including 24 each of African-Americans from Jackson, Mississippi; European-Americans from Rochester, Minnesota; and Europeans from North Karelia, Finland. This sequence survey detected 21 SNPs and 1 multiallelic indel, 14 of which had not been previously reported. Alleles varied in relative frequency among the populations, and 10 sites were polymorphic in only a single population sample. Oligonucleotide ligation assays (OLA) were developed for 20 of these sites (omitting the indel and a closely-linked SNP). These were then scored in 2179 individuals sampled from the same three populations (n = 843, 884, and 452, respectively). Relative allele frequencies were generally consistent with estimates from the core sample, although variation was found in some populations in the larger sample at SNPs that were monomorphic in the corresponding smaller core sample. Site variation in the larger samples showed no systematic deviation from Hardy-Weinberg expectation. The large OLA sample clearly showed that variation in many, but not all, of OLA-typed SNPs is significantly correlated with the classical protein-coding variants, implying that there may be important substructure within the classical ɛ2, ɛ3, and ɛ4 alleles. Comparison of the levels and patterns of polymorphism in the core samples with those estimated for the OLA-typed samples shows how nucleotide diversity is underestimated when

  20. The removal of cholesterol from aortic smooth muscle cells in culture and Landschutz ascites cells by fractions of human high-density apolipoprotein.

    PubMed

    Stein, Y; Glangeaud, M C; Fainaru, M; Stein, O

    1975-01-24

    Ascites cells were labeled by intraperitoneal injection of [3H]cholesterol and aortic smooth muscle cells by addition of [3H]cholesterol to the serum component of the culture medium. The release of cholesterol from cells into a serum-free medium supplemented with the various "acceptors" was studied using ascites cells in suspension and aortic smooth muscle cells in a multilayer culture. Unfractionated human high-density apolipoprotein was somewhat more effective in the removal of labeled cellular free cholesterol, in both cell types, than apolipoprotein derived from rat high-density lipoprotein. Following separation of human high-density apolipoprotein into four fractions by Sephadex chromatography, the effect of each fraction on the removal of cellular cholesterol from ascites cells was studied. The individual fractions had a lower capacity for cholesterol removal than the original unfractionated high-density apolipoprotein and the lowest activity was detected in Fraction II which comprised 75% of the total apolipoprotein. The effectiveness to remove cholesterol could be restored to all the fractions, as well as enhanced, by addition of sonicated suspensions of lecithin or sphingomyelin, which by themselves promoted a more limited removal of cellular cholesterol. Negatively stained preparations of mixtures of the four fractions and sonicated dispersion of lecithin were shown to consist of vesicles and discs of various sizes. Addition of the apolipoprotein fractions (especially Fractions II and IV) to sonicated dispersion of sphingomyelin resulted in a pronounced formation of discs which showed a high tendency towards stack formation. Mixtures of Fraction II and lecithin or sphingomyelin were effective in the release of cellular cholesterol from multilayers of aortic smooth muscle cells in culture. These results show the feasibility of net removal of cholesterol from cells which grow in a form resembling a tissue and thus provide a model to study the role of

  1. C/EBP-β Is Differentially Affected by PPARα Agonists Fenofibric Acid and GW7647, But Does Not Change Apolipoprotein A-I Production During ER-Stress and Inflammation.

    PubMed

    van der Krieken, Sophie E; Popeijus, Herman E; Konings, Maurice; Dullens, Stefan P J; Mensink, Ronald P; Plat, Jogchum

    2017-04-01

    Increasing apolipoproteinA-I (apoA-I) production may be anti-atherogenic. Thus, there is a need to identify regulatory factors involved. Transcription of apoA-I involves peroxisome-proliferator-activated-receptor-alpha (PPARα) activation, but endoplasmic reticulum (ER) -stress and inflammation also influence apoA-I production. To unravel why PPARα agonist GW7647 increased apoA-I production compared to PPARα agonist fenofibric acid (FeAc) in human hepatocellular carcinoma (HepG2) and colorectal adenocarcinoma (CaCo-2) cells, gene expression profiles were compared. Microarray analyses suggested CCAAT/enhancer-binding-protein-beta (C/EBP-β) involvement in the FeAc condition. Therefore, C/EBP-β silencing and isoform-specific overexpression experiments were performed under ER-stressed, inflammatory and non-inflammatory conditions. mRNA expression of C/EBP-β, ATF3, NF-IL3 and GDF15 were upregulated by FeAc compared to GW7647 in both cell lines, while DDIT3 and DDIT4 mRNA were only upregulated in HepG2 cells. This ER-stress related signature was associated with decreased apoA-I secretion. After ER-stress induction by thapsigargin or FeAc addition, intracellular apoA-I concentrations decreased, while ER-stress marker expression (CHOP, XBP1s, C/EBP-β) increased. Cytokine addition increased intracellular C/EBP-β levels and lowered apoA-I concentrations. Although a C/EBP binding place is present in the apoA-I promoter, C/EBP-β silencing or isoform-specific overexpression did not affect apoA-I production in inflammatory, non-inflammatory and ER-stressed conditions. Therefore, C/EBP-β is not a target to influence hepatic apoA-I production. J. Cell. Biochem. 118: 754-763, 2017. © 2016 Wiley Periodicals, Inc.

  2. High- and low-temperature unfolding of human high-density apolipoprotein A-2.

    PubMed

    Gursky, O; Atkinson, D

    1996-09-01

    Human plasma apolipoprotein A-2 (apoA-2) is the second major protein of the high-density lipoproteins that mediate the transport and metabolism of cholesterol. Using CD spectroscopy and differential scanning calorimetry, we demonstrate that the structure of lipid-free apoA-2 in neutral low-salt solutions is most stable at approximately 25 degrees C and unfolds reversibly both upon heating and cooling from 25 degrees C. High-temperature unfolding of apoA-2, monitored by far-UV CD, extends from 25-85 degrees C with midpoint Th = 56 +/- 2 degrees C and vant Hoff's enthalpy delta H(Th) = 17 +/- 2 kcal/mol that is substantially lower than the expected enthalpy of melting of the alpha-helical structure. This suggests low-cooperativity apoA-2 unfolding. The apparent free energy of apoA-2 stabilization inferred from the CD analysis of the thermal unfolding, delta G(app)(25 degrees) = 0.82 +/- 0.15 kcal/mol, agrees with the value determined from chemical denaturation. Enhanced low-temperature stability of apoA-2 observed upon increase in Na2HPO4 concentration from 0.3 mM to 50 mM or addition of 10% glycerol may be linked to reduced water activity. The close proximity of the heat and cold unfolding transitions, that is consistent with low delta G(app)(25 degrees), indicates that lipid-free apoA-2 has a substantial hydrophobic core but is only marginally stable under near-physiological solvent conditions. This suggests that in vivo apoA-2 transfer is unlikely to proceed via the lipid-free state. Low delta H(Th) and low apparent delta Cp approximately 0.52 kcal/mol.K inferred from the far-UV CD analysis of apoA-2 unfolding, and absence of tertiary packing interactions involving Tyr groups suggested by near-UV CD, are consistent with a molten globular-like state of lipid-free apoA-2.

  3. Human plasma lipid modulation in schistosomiasis mansoni depends on apolipoprotein E polymorphism.

    PubMed

    Martins da Fonseca, Caíque Silveira; Pimenta Filho, Adenor Almeida; dos Santos, Bianka Santana; da Silva, César Augusto; Domingues, Ana Lúcia Coutinho; Owen, James Stuart; Lima, Vera Lúcia de Menezes

    2014-01-01

    Schistosomiasis mansoni is a parasitic liver disease, which causes several metabolic disturbances. Here, we evaluate the influence of Apolipoprotein E (APOE) gene polymorphism, a known modulator of lipid metabolism, on plasma lipid levels in patients with hepatosplenic schistosomiasis. Blood samples were used for APOE genotyping and to measure total cholesterol (TC), LDL-C, HDL-C and triglycerides. Schistosomiasis patients had reduced TC, LDL-C and triglycerides (25%, 38% and 32% lower, respectively; P<0.0001) compared to control individuals, whereas HDL-C was increased (10% higher; P = 0.0136). Frequency of the common alleles, ε2, ε3 and ε4, was similar (P = 0.3568) between controls (n = 108) and patients (n = 84), implying that APOE genotype did not affect susceptibility to the advanced stage of schistosomiasis. Nevertheless, while patient TC and LDL-C levels were significantly reduced for each allele (except TC in ε2 patients), changes in HDL-C and triglycerides were noted only for the less common ε2 and ε4 alleles. The most striking finding, however, was that accepted regulation of plasma lipid levels by APOE genotype was disrupted by schistosomiasis. Thus, while ε2 controls had higher TC and LDL-C than ε3 carriers, these parameters were lower in ε2 versus ε3 patients. Similarly, the inverse relationship of TG levels in controls (ε2>ε3>ε4) was absent in patients (ε2 or ε4>ε3), and the increase in HDL-C of ε2 or ε4 patients compared to ε3 patients was not seen in the control groups. We confirm that human schistosomiasis causes dyslipidemia and report for the first time that certain changes in plasma lipid and lipoprotein levels depend on APOE gene polymorphism. Importantly, we also concluded that S. mansoni disrupts the expected regulation of plasma lipids by the different ApoE isoforms. This finding suggests ways to identify new metabolic pathways affected by schistosomiasis and also potential molecular targets to treat

  4. Role of Conserved Proline Residues in Human Apolipoprotein A-IV Structure and Function*

    PubMed Central

    Deng, Xiaodi; Walker, Ryan G.; Morris, Jamie; Davidson, W. Sean; Thompson, Thomas B.

    2015-01-01

    Apolipoprotein (apo)A-IV is a lipid emulsifying protein linked to a range of protective roles in obesity, diabetes, and cardiovascular disease. It exists in several states in plasma including lipid-bound in HDL and chylomicrons and as monomeric and dimeric lipid-free/poor forms. Our recent x-ray crystal structure of the central domain of apoA-IV shows that it adopts an elongated helical structure that dimerizes via two long reciprocating helices. A striking feature is the alignment of conserved proline residues across the dimer interface. We speculated that this plays important roles in the structure of the lipid-free protein and its ability to bind lipid. Here we show that the systematic conversion of these prolines to alanine increased the thermodynamic stability of apoA-IV and its propensity to oligomerize. Despite the structural stabilization, we noted an increase in the ability to bind and reorganize lipids and to promote cholesterol efflux from cells. The novel properties of these mutants allowed us to isolate the first trimeric form of an exchangeable apolipoprotein and characterize it by small-angle x-ray scattering and chemical cross-linking. The results suggest that the reciprocating helix interaction is a common feature of all apoA-IV oligomers. We propose a model of how self-association of apoA-IV can result in spherical lipoprotein particles, a model that may have broader applications to other exchangeable apolipoprotein family members. PMID:25733664

  5. Apolipoprotein D is the major protein component in cyst fluid from women with human breast gross cystic disease.

    PubMed Central

    Balbín, M; Freije, J M; Fueyo, A; Sánchez, L M; López-Otín, C

    1990-01-01

    GCDFP(gross-cystic-disease-fluid protein)-24, a progesterone-binding protein present in large amounts in cyst fluid from human breast gross cystic disease, was purified in a one-step procedure by size-exclusion h.p.l.c. Peptide fragments obtained by trypsin digestion of the intact protein were purified by reverse-phase h.p.l.c. and analysed for their amino acid composition and subjected to automated Edman degradation. A search of the National Biomedical Research Foundation Data Bank revealed that all the sequenced tryptic peptides from protein GCDFP-24 matched perfectly with regions present in the amino acid sequence determined for human apolipoprotein D. Additional data on N-terminal sequence of the unblocked proteins, carbohydrate-attachment sites, amino acid composition and molecular-mass estimations supported the identity between both molecules. On the basis of this identity a possible role of apolipoprotein D in progesterone transport is proposed. Images Fig. 2. PMID:2244881

  6. Effect of human apolipoprotein E genotype on the pathogenesis of experimental ocular HSV-1

    PubMed Central

    Bhattacharjee, Partha S.; Neumann, Donna M.; Foster, Timothy P.; Bouhanik, Sadallah; Clement, Christian; Vinay, Dass; Thompson, Hilary W.; Hill, James M.

    2008-01-01

    The isoform-specific role of human apolipoprotein E (apoE) has been assessed in a mouse model of ocular herpes. Female, age-matched transgenic mice knocked-in for the human allele apoE3 or apoE4 and their parent C57Bl/6 mice were inoculated corneally with HSV-1 strain KOS. Ocular HSV-1 pathogenesis was monitored through viral replication and clinical progression of stromal opacity and neovascularization by slit-lamp examination. Establishment of latency was determined by analysis of HSV-1 DNA (copy number) by specific real-time PCR in the cornea, trigeminal ganglia (TG), and brain. Representative groups of transgenic mice were sacrificed for the analysis of gene expression of vascular endothelial growth factor (VEGF) by reverse-transcription PCR, and apoE expression by Western blot analysis. At 6 days post-infection (P.I.), the ocular infectious HSV-1 titer was significantly higher (p < 0.05) in apoE4 mice compared with apoE3 and C57Bl/6 mice. Corneal neovascularization in apoE4 mice was significantly higher (p < 0.05) than apoE3 and C57Bl/6 mice. The onset of corneal opacity in apoE4 mice was accelerated during days 9--11 P.I.; however, no significant difference in severity was seen on P.I. days 15 and beyond. At 28 days P.I., infected mice of all genotypes had no significant differences in copy numbers (range 0--15) of HSV-1 DNA in their corneas, indicating that HSV-1 DNA copy numbers in cornea are independent of apoE isoform regulation. At 28 days P.I., both apoE4 and C57Bl/6 mice had a significantly higher (p = 0.001) number of copies of HSV-1 DNA in TG compared with apoE3. ApoE4 mice also had significantly higher (p = 0.001) copies of HSV-1 DNA in their TGs compared with C57Bl/6 mice. In brain, both apoE4 and C57Bl/6 mice had significantly higher numbers (p ≤ 0.03) of copies of HSV-1 DNA compared with apoE3 mice. However, the number of HSV-1 DNA copies in the brain of C57Bl/6 mice was not significantly different than that of apoE4 (p = 0.1). Comparative

  7. 18O proteomics reveal increased Human Apolipoprotein CIII in Hispanic HIV-1 positive women with HAART that use cocaine

    PubMed Central

    Zenón, Frances; Jorge, Inmaculada; Cruz, Ailed; Suarez, Erick; Segarra, Annabell C.; Vázquez, Jesús; Meléndez, Loyda M.; Serrano, Horacio

    2016-01-01

    Purpose Drug abuse is a major risk factor in the development and progression of HIV-1. This study defines the alterations in the plasma proteome of HIV-1 infected women that use cocaine. Experimental Design Plasma samples from 12 HIV-seropositive Hispanic women under antiretroviral therapy were selected for this study. Six sample pairs were matched between non-drug users and cocaine users. After IgG and albumin depletion, SDS-PAGE, and in-gel digestion, peptides from non-drug users and cocaine users were labeled with 16O and 18O respectively and subjected to LC-MS/MS and quantitation using Proteome Discover and QuiXoT softwares and validated by ELISA. Results A total of 1,015 proteins were identified at 1% FDR. Statistical analyses revealed 13 proteins with significant changes between the two groups, cocaine and non-cocaine users (p<0.05). The great majority pertained to protection defense function and the rest pertained to transport, homeostatic, regulation, and binding of ligands. Apolipoprotein CIII was increased in plasma of HIV+ Hispanic women positive for cocaine compared to HIV+ non-drug users (p<0.05). Conclusions and clinical relevance Increased human Apolipoprotein CIII warrants that these patients be carefully monitored to avoid the increased risk of cardiovascular events associated with HIV, HAART and cocaine use. PMID:26255783

  8. 18O proteomics reveal increased human apolipoprotein CIII in Hispanic HIV-1+ women with HAART that use cocaine.

    PubMed

    Zenón, Frances; Jorge, Inmaculada; Cruz, Ailed; Suárez, Erick; Segarra, Annabell C; Vázquez, Jesús; Meléndez, Loyda M; Serrano, Horacio

    2016-02-01

    Drug abuse is a major risk factor in the development and progression of HIV-1. This study defines the alterations in the plasma proteome of HIV-1-infected women that use cocaine. Plasma samples from 12 HIV-seropositive Hispanic women under antiretroviral therapy were selected for this study. Six sample pairs were matched between nondrug users and cocaine users. After IgG and albumin depletion, SDS-PAGE, and in-gel digestion, peptides from nondrug users and cocaine users were labeled with (16) O and (18) O, respectively, and subjected to LC-MS/MS and quantitation using Proteome Discover and QuiXoT softwares and validated by ELISA. A total of 1015 proteins were identified at 1% false discovery rates (FDR). Statistical analyses revealed 13 proteins with significant changes between the two groups, cocaine and noncocaine users (p < 0.05). The great majority pertained to protection defense function and the rest pertained to transport, homeostatic, regulation, and binding of ligands. Apolipoprotein CIII was increased in plasma of HIV+ Hispanic women positive for cocaine compared to HIV+ nondrug users (p ≤ 0.05). Increased human apolipoprotein CIII warrants that these patients be carefully monitored to avoid the increased risk of cardiovascular events associated with HIV, HAART, and cocaine use. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Fractionation of apolipoproteins from human serum very low density lipoproteins by chromatofocusing.

    PubMed

    Jauhiainen, M

    1982-01-01

    1. A pooled serum from several pregnant women was used as a source of VLDL 2. VLDL and if needed other lipoproteins were fractionated by sequential flotation. 3. Lipoproteins were delipidated and lipid-free VLDL apolipoproteins were fractionated by a new chromatofocusing technique. 4. Chromatofocusing column run yielded 7 peak protein fractions and the corresponding pI values were: 6.8, 6.6, 5.7, 5.5, 5.2, 4.8 and 4.4. 5. Polyacrylamide slab gel electrophoresis of the chromatofocusing protein peaks indicated that they are different having dissimilar Rf values in urea-SDS containing slabs.

  10. NMR structure of human apolipoprotein C-II in the presence of sodium dodecyl sulfate.

    PubMed

    MacRaild, C A; Hatters, D M; Howlett, G J; Gooley, P R

    2001-05-08

    The structure and protein-detergent interactions of apolipoprotein C-II (apoC-II) in the presence of SDS micelles have been investigated using circular dichroism and heteronuclear NMR techniques applied to (15)N-labeled protein. Micellar SDS, a commonly used mimetic of the lipoprotein surface, inhibits the aggregation of apoC-II and induces a stable structure containing approximately 60% alpha-helix as determined by circular dichroism. NMR reveals the first 12 residues of apoC-II to be structurally heterogeneous and largely disordered, with the rest of the protein forming a predominantly helical structure. Three regions of helical conformation, residues 16-36, 50-56, and 63-77, are well-defined by NMR-derived constraints, with the intervening regions showing more loosely defined helical conformation. The structure of apoC-II is compared to that determined for other apolipoproteins in a similar environment. Our results shed light on the lipid interactions of apoC-II and its mechanism of lipoprotein lipase activation.

  11. Inhibition of radical reaction of apolipoprotein B-100 and alpha-tocopherol in human plasma by green tea catechins.

    PubMed

    Hashimoto, R; Yaita, M; Tanaka, K; Hara, Y; Kojo, S

    2000-12-01

    (-)-epicatechin (EC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECg), (-)-epigallocatechin gallate (EGCg), and Trolox inhibited the decreases of apolipoprotein B-100 (apoB) and alpha-tocopherol in a radical reaction of human plasma initiated by Cu(2+). The concentrations of EC, EGC, ECg, EGCg, and Trolox for 50% inhibition (IC50) of apoB fragmentation were 39.1, 42.2, 14.6, 21.3, and 36.2 microM, respectively. Similar IC50 values were observed for alpha-tocopherol consumption, indicating the close relationship between apoB fragmentation and alpha-tocopherol consumption. These results demonstrate that tea catechins serve as an effective antioxidant in plasma and that the gallate group has a strong antioxidative activity.

  12. Biosynthesis of high density lipoprotein by chicken liver: intracellular transport and proteolytic processing of nascent apolipoprotein A-1

    PubMed Central

    1985-01-01

    To study the in vivo processing and secretion of Apolipoprotein A-I (Apo A-I), young chickens were administered individual L-[3H]amino acids intravenously and the time of intracellular transport of nascent Apo A-I from rough endoplasmic reticulum (RER) to the Golgi apparatus was measured. Within 3 to 9 min there was maximal incorporation of radioactivity into Apo A-I in both the RER and the Golgi cell fractions. By contrast, the majority of radioactive albumin was also present in the RER by 3 to 9 min, but did not reach peak amounts in the Golgi fraction until 9 to 25 min. Both radioactive Apo A-I and albumin appeared in the blood at about the same time (between 20 and 30 min). NH2-terminal amino acid sequence analysis of nascent intracellular Apo A-I showed that it contains a pro-hexapeptide extension identical to that of human Apo A-I. After 30 min of administration of radioactive amino acids radioactive Apo A-I was isolated by immunoprecipitation from the liver and serum. NH2-terminal sequence analysis of 20 amino acids indicated that chicken liver contained an equal mixture of nascent pro-Apo A-I and fully processed Apo A-I, whereas the serum only contained processed Apo A-I. Further studies showed that the RER only contained pro-Apo A-I, whereas a mixture of pro-Apo A-I and processed Apo A-I was found in the Golgi complex. These results indicate that, in chicken hepatocytes, there is a more rapid transport of Apo A-I than of albumin from the RER to the Golgi cell fractions, and that Apo A-I remains in the Golgi apparatus for a longer period of time before it is secreted into the blood. In addition these studies show that the in vivo proteolytic processing of chicken pro-Apo A-I to Apo A-I occurs in the Golgi cell fractions. PMID:3930506

  13. Apolipoprotein distribution in human lipoproteins separated by polyacrylamide gradient gel electrophoresis.

    PubMed

    Vézina, C A; Milne, R W; Weech, P K; Marcel, Y L

    1988-05-01

    The heterogeneity of serum lipoproteins (excluding very low density (VLDL) and intermediate density (IDL) lipoproteins) and that of lipoproteins secreted by HepG2 cells has been studied by immunoblot analysis of the apolipoprotein composition of the particles separated by polyacrylamide gradient gel electrophoresis (GGE) under nondenaturing conditions. The reactions of antibodies to apoA-I, apoA-II, apoE, apoB, apoD, and apoA-IV have revealed discrete bands of particles which differ widely in size and apolipoprotein composition. GGE of native serum lipoproteins demonstrated that apoA-II is present in lipoproteins of limited size heterogeneity (apparent molecular mass 345,000 to 305,000) and that apoB is present in low density lipoproteins (LDL) and absent from all smaller or denser lipoproteins. In contrast, serum apoA-I, E, D, and A-IV are present in very heterogeneous particles. Serum apoA-I is present mainly in particles of 305 to 130 kDa where it is associated with apoA-II, and in decreasing order of immunoreactivity in particles of 130-90 kDa, 56 kDa, 815-345 kDa, and finally within the size range of LDL, all regions where there is little detectable apoA-II. Serum apoE is present in three defined fractions, one within the size range of LDL, one containing heterogeneous particles between 640 and 345 kDa, and one defined fraction at 96 kDa. Serum apoD is also present in three defined fractions, one comigrating with LDL, one containing heterogeneous particles between 390 and 150 kDa, and one band on the migration front. Most of serum apoA-IV is contained in a band comigrating with albumin. GGE of centrifugally prepared LDL shows the presence of apoB, apoE, and apoD, but not that of apoA-I. However, the particles containing apoA-I, which, in serum, migrated within the LDL size range and as bands of 815 to 345 kDa, were recovered upon centrifugation in the d greater than 1.21 g/ml fraction. GGE of high density lipoproteins (HDL) indicated that most of apoA-I, A

  14. Brazil nut ingestion increased plasma selenium but had minimal effects on lipids, apolipoproteins, and high-density lipoprotein function in human subjects.

    PubMed

    Strunz, Célia C; Oliveira, Tatiane V; Vinagre, Juliana C M; Lima, Adriana; Cozzolino, Silvia; Maranhão, Raul C

    2008-03-01

    The Brazil nut (Bertholletia excelsa) of the Amazon region is consumed worldwide. It is rich in both monounsaturated fatty acids and polyunsaturated fatty acids and is known for its high selenium content. This study tested the hypothesis whether the consumption of this nut could affect the plasma lipids and apolipoproteins and some functional properties of the antiatherogenic high-density lipoprotein (HDL). Fifteen normolipidemic subjects aged 27.3 +/- 3.9 years and with body mass index of 23.8 +/- 2.8 kg/m(2) consumed 45 g of Brazil nuts per day during a 15-day period. On days 0 and 15, blood was collected for biochemical analysis, determination of HDL particle size, paraoxonase 1 activity, and lipid transfer from a lipoprotein-like nanoparticle to the HDL fraction. Brazil nut ingestion did not alter HDL, low-density lipoprotein cholesterol, triacylglycerols, apolipoprotein A-I, or apolipoprotein B concentrations. HDL particle diameter and the activity of antioxidative paraoxonase 1, mostly found in the HDL fraction, were also unaffected. Supplementation increased the reception of cholesteryl esters (P < .05) by the HDL yet did not alter the reception of phospholipids, free cholesterol, or triacylglycerols. As expected, plasma selenium was significantly increased. However, the consumption of Brazil nuts for short duration by normolipidemic subjects in comparable amounts to those tested for other nuts did not alter serum lipid profile. The only alteration in HDL function was the increase in cholesteryl ester transfer. This latter finding may be beneficial because it would improve the nonatherogenic reverse cholesterol transport pathway.

  15. Mechanism of lipid lowering in mice expressing human apolipoprotein A5

    SciTech Connect

    Fruchart-Najib, Jamila; Bauge, Eric; Niculescu, Loredan-Stefan; Pham, Tatiana; Thomas, Benoit; Rommens, Corinne; Majd, Zouher; Brewer, Bryan; Rubin, Edward M.; Pennacchio, Len A.; Fruchart, Jean-Charles

    2004-01-15

    Recently, we reported that apoAV plays key role in triglycerides lowering. Here, we attempted to determine the mechanism underlying this hypotriglyceridemic effect. We showed that triglyceride turnover is faster in hAPOA5 transgenic compared to wild type mice. Moreover, both apoB and apoCIII are decreased and LPL activity is increased in postheparin plasma of hAPOA5 transgenic mice. These data suggest a decrease in size and number of VLDL. To further investigate the mechanism of hAPOA5 in hyperlipidemic background, we intercrossed hAPOA5 and hAPOC3 transgenic mice. The effect resulted in a marked decreased of VLDL triglyceride, cholesterol, apolipoproteins B and CIII. In postprandial state, the triglyceride response is abolished in hAPOA5 transgenic mice. We demonstrated that in response to the fat load in hAPOA5XhAPOC3 mice, apoAV shifted from HDL to VLDL, probably to limit the elevation of triglycerides. In vitro, apoAV activates lipoprotein lipase. However, apoAV does not interact with LPL but interacts physically with apoCIII. This interaction does not seem to displace apoCIII from VLDL but may induce conformational change in apoCIII and consequently change in its function leading the activation of lipoprotein lipase.

  16. Human apolipoprotein E ɛ4 expression impairs cerebral vascularization and blood–brain barrier function in mice

    PubMed Central

    Alata, Wael; Ye, Yue; St-Amour, Isabelle; Vandal, Milène; Calon, Frédéric

    2015-01-01

    Human apolipoprotein E (APOE) exists in three isoforms ɛ2, ɛ3, and ɛ4, of which APOE4 is the main genetic risk factor of Alzheimer's disease (AD). As cerebrovascular defects are associated with AD, we tested whether APOE genotype has an impact on the integrity and function of the blood–brain barrier (BBB) in human APOE-targeted replacement mice. Using the quantitative in situ brain perfusion technique, we first found lower (13.0% and 17.0%) brain transport coefficient (Clup) of [3H]-diazepam in APOE4 mice at 4 and 12 months, compared with APOE2 and APOE3 mice, reflecting a decrease in cerebral vascularization. Accordingly, results from immunohistofluorescence experiments revealed a structurally reduced cerebral vascularization (26% and 38%) and thinner basement membranes (30% and 35%) in 12-month-old APOE4 mice compared with APOE2 and APOE3 mice, suggesting vascular atrophy. In addition, APOE4 mice displayed a 29% reduction in [3H]-d-glucose transport through the BBB compared with APOE2 mice without significant changes in the expression of its transporter GLUT1 in brain capillaries. However, an increase of 41.3% of receptor for advanced glycation end products (RAGE) was found in brain capillaries of 12-month-old APOE4 mice. In conclusion, profound divergences were observed between APOE genotypes at the cerebrovascular interface, suggesting that APOE4-induced BBB anomalies may contribute to AD development. PMID:25335802

  17. Cytokines decrease apolipoprotein accumulation in medium from Hep G2 cells.

    PubMed

    Ettinger, W H; Varma, V K; Sorci-Thomas, M; Parks, J S; Sigmon, R C; Smith, T K; Verdery, R B

    1994-01-01

    Cytokines, important biochemical mediators of inflammation, cause a rapid fall in the plasma concentration of cholesterol in vivo. One mechanism by which cytokines may cause acquired hypocholesterolemia is by decreasing the hepatic synthesis and secretion of apolipoproteins. To test this hypothesis, we incubated Hep G2 cells with human recombinant tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. Each of the cytokines resulted in a dose-related reduction in the concentrations of apolipoprotein (apo) A-I, apoB, and lecithin:cholesterol acyltransferase (LCAT) activity in the medium after 24 hours of incubation. The effect of cytokines on apolipoprotein accumulation was not affected by preincubation of Hep G2 cells with fatty acids. Cytokines decreased the concentration of cellular apoA-I mRNA in a dose-related fashion but did not affect cellular concentrations of apoB mRNA. The concentrations of triglyceride and cholesterol were also reduced in the medium of cells incubated with cytokines. Total cell sterol synthesis rates were calculated by [14C]acetate incorporation. Cells incubated with interleukin-6 had a 31% increase in sterol synthesis rate but a 41% decrease in sterol secretion. These data suggest that these cytokines can decrease the hepatic synthesis and/or secretion of apolipoproteins and that this may explain, in part, the acquired hypocholesterolemia seen during acute and chronic inflammation.

  18. Intravenously Injected Human Apolipoprotein A‐I Rapidly Enters the Central Nervous System via the Choroid Plexus

    PubMed Central

    Stukas, Sophie; Robert, Jerome; Lee, Michael; Kulic, Iva; Carr, Michael; Tourigny, Katherine; Fan, Jianjia; Namjoshi, Dhananjay; Lemke, Kalistyne; DeValle, Nicole; Chan, Jeniffer; Wilson, Tammy; Wilkinson, Anna; Chapanian, Rafi; Kizhakkedathu, Jayachandran N.; Cirrito, John R.; Oda, Michael N.; Wellington, Cheryl L.

    2014-01-01

    Background Brain lipoprotein metabolism is dependent on lipoprotein particles that resemble plasma high‐density lipoproteins but that contain apolipoprotein (apo) E rather than apoA‐I as their primary protein component. Astrocytes and microglia secrete apoE but not apoA‐I; however, apoA‐I is detectable in both cerebrospinal fluid and brain tissue lysates. The route by which plasma apoA‐I enters the central nervous system is unknown. Methods and Results Steady‐state levels of murine apoA‐I in cerebrospinal fluid and interstitial fluid are 0.664 and 0.120 μg/mL, respectively, whereas brain tissue apoA‐I is ≈10% to 15% of its levels in liver. Recombinant, fluorescently tagged human apoA‐I injected intravenously into mice localizes to the choroid plexus within 30 minutes and accumulates in a saturable, dose‐dependent manner in the brain. Recombinant, fluorescently tagged human apoA‐I accumulates in the brain for 2 hours, after which it is eliminated with a half‐life of 10.3 hours. In vitro, human apoA‐I is specifically bound, internalized, and transported across confluent monolayers of primary human choroid plexus epithelial cells and brain microvascular endothelial cells. Conclusions Following intravenous injection, recombinant human apoA‐I rapidly localizes predominantly to the choroid plexus. Because apoA‐I mRNA is undetectable in murine brain, our results suggest that plasma apoA‐I, which is secreted from the liver and intestine, gains access to the central nervous system primarily by crossing the blood–cerebrospinal fluid barrier via specific cellular mediated transport, although transport across the blood–brain barrier may also contribute to a lesser extent. PMID:25392541

  19. P-selectin expressed by a human SELP transgene is atherogenic in apolipoprotein E-deficient mice

    PubMed Central

    Zhang, Nan; Liu, Zhenghui; Yao, Longbiao; Mehta-D’souza, Padmaja; McEver, Rodger P.

    2016-01-01

    Objective During inflammation, P-selectin expressed on activated endothelial cells and platelets mediates rolling adhesion of leukocytes. Atherosclerosis-prone mice crossed with P-selectin-deficient (Selp−/−) mice develop smaller lesions. Cytokines such as tumor necrosis factor-α increase Selp transcripts and augment atherosclerosis in mice. However, they decrease SELP transcripts in humans, challenging assumptions that human P-selectin is atherogenic. We used mice expressing a human SELP transgene to examine the atherogenic role of P-selectin. Approach and results We crossed apolipoprotein E-deficient (Apoe−/−) mice with Selp−/− mice and/or transgenic mice expressing the entire human SELP gene (TgSELP+/−). Aortas developed larger, macrophage-rich atheromas in Apoe−/−Selp−/−TgSELP+/− mice than in Apoe−/−Selp−/− mice after 8 or 16 weeks on a Western diet. Confocal microscopy of Apoe−/−Selp−/−TgSELP+/− aortas revealed staining for human P-selectin in endothelial cells overlying atheromas, but not in lesional macrophages. We also observed staining for human P-selectin in aortic endothelial cells of 3–4-week-old Apoe−/−Selp−/−TgSELP+/− weanlings before atheromas developed. Furthermore, human SELP transcripts were ~3-fold higher in aortas of Apoe−/−Selp+/−TgSELP+/− weanlings than in Selp+/−TgSELP+/− weanlings, whereas murine Selp and Sele transcripts were equivalent in weanlings of both genotypes. Human SELP transcripts in aortas of Apoe−/−Selp+/−TgSELP+/− mice remained nearly constant during 16 weeks on a Western diet, whereas murine Selp and Sele transcripts progressively increased. Bone marrow transplantation in Apoe−/−Selp−/− and Apoe−/−Selp−/−TgSELP+/− mice demonstrated that both platelets and endothelial cells must express human P-selectin to promote atherogenesis. Conclusions P-selectin expressed by human SELP is atherogenic in Apoe−/− mice, suggesting that P

  20. Susceptibility of Mice to Trypanosoma evansi Treated with Human Plasma Containing Different Concentrations of Apolipoprotein L-1

    PubMed Central

    Fanfa, Vinicius R.; Otto, Mateus A.; Gressler, Lucas T.; Tavares, Kaio C.S.; Lazzarotto, Cícera R.; Tonin, Alexandre A.; Miletti, Luiz C.; Duarte, Marta M.M.F.; Monteiro, Silvia G.

    2011-01-01

    The aim of this study was to test the susceptibility of mice to Trypanosoma evansi treated with human plasma containing different concentrations of apolipoprotein L-1 (APOL1). For this experiment, a strain of T. evansi and human plasma (plasmas 1, 2, and 3) from 3 adult males clinically healthy were used. In vivo test used 50 mice divided in 5 groups (A to E) with 10 animals in each group. Animals of groups B to E were infected, and then treated with 0.2 ml of human plasma in the following outline: negative control (A), positive control (B), treatment with plasma 1 (C), treatment with plasma 2 (D), and treatment with plasma 3 (E). Mice treated with human plasma showed an increase in longevity of 40.9±0.3 (C), 20±9.0 (D) and 35.6±9.3 (E) days compared to the control group (B) which was 4.3±0.5 days. The number of surviving mice and free of the parasite (blood smear and PCR negative) at the end of the experiment was 90%, 0%, and 60% for groups C, D, and E, respectively. The quantification of APOL1 was performed due to the large difference in the treatments that differed in the source plasma. In plasmas 1, 2, and 3 was detected the concentration of 194, 99, and 115 mg/dl of APOL1, respectively. However, we believe that this difference in the treatment efficiency is related to the level of APOL1 in plasmas. PMID:22355213

  1. Inhibition of transglutaminase 2 reduces efferocytosis in human macrophages: Role of CD14 and SR-AI receptors.

    PubMed

    Eligini, S; Fiorelli, S; Tremoli, E; Colli, S

    2016-10-01

    Transglutaminase 2 (TGM2), a member of the transglutaminase family of enzymes, is a multifunctional protein involved in numerous events spanning from cell differentiation, to signal transduction, apoptosis, and wound healing. It is expressed in a variety of cells, macrophages included. Macrophage TGM2 promotes the clearance of apoptotic cells (efferocytosis) and emerging evidence suggests that defective efferocytosis contributes to the consequences of inflammation-associated diseases, including atherosclerotic lesion progression and its sequelae. Of interest, active TGM2 identified in human atherosclerotic lesions plays critical roles in plaque stability through effects on matrix cross-linking and TGFβ activity. This study explores the mechanisms by which TGM2 controls efferocytosis in human macrophages. Herein we show that TGM2 increases progressively during monocyte differentiation towards macrophages and controls their efferocytic potential as well as morphology and viability. Two experimental approaches that took advantage of the inhibition of TGM2 activity and protein silencing give proof that TGM2 reduction significantly impairs macrophage efferocytosis. Among the mechanisms involved we highlighted a role of the receptors CD14 and SR-AI whose levels were markedly reduced by TGM2 inhibition. Conversely, CD36 receptor and αvβ3 integrin levels were not influenced. Of note, lipid accumulation and IL-10 secretion were reduced in macrophages displaying defective efferocytosis. Overall, our data define a crucial role of TGM2 activity during macrophage differentiation via mechanisms involving CD14 and SR-AI receptors and show that TGM2 inhibition triggers a pro-inflammatory phenotype. Copyright © 2016 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

  2. Human recombinant apolipoprotein E redirects lipopolysaccharide from Kupffer cells to liver parenchymal cells in rats In vivo.

    PubMed Central

    Rensen, P C; Oosten, M; Bilt, E; Eck, M; Kuiper, J; Berkel, T J

    1997-01-01

    Chylomicrons have been shown to protect mice and rats against a lethal dose of lipopolysaccharide and may serve as a therapeutic means to protect against endotoxemia. However, the requisite of isolation from human lymph hampers pharmaceutical application. Recently, we developed recombinant chylomicrons from commercially available lipids and human recombinant apolipoprotein E. The current study explored the effectiveness of these apoE-enriched emulsions in redirecting LPS from Kupffer cells to liver parenchymal cells. Upon injection into rats, 125I-LPS rapidly and specifically associated with the liver (64.3+/-3.1% of the injected dose) and spleen (4.1+/-0.7%). The uptake of LPS by the spleen was four- to fivefold reduced upon incubation with the apoE-enriched emulsion or free apoE (P < 0.0001), but not with emulsion alone or Lipofundin. Within the liver, 125I-LPS mainly associated with Kupffer cells. The uptake by Kupffer cells was eight- to ninefold reduced by the apoE-enriched emulsion or apoE alone (P < 0.01), and a 19.6-fold increased uptake ratio by liver parenchymal cells over Kupffer cells was observed. The emulsion without apoE had no effect on the in vivo kinetics of LPS. LPS interacted selectively with the apoE moiety of the recombinant chylomicron. Emulsion-associated and free apoE bound approximately two molecules of LPS, possibly by its exposed hydrophilic domain involving arginine residues. We anticipate that the protecting effect of endogenous chylomicrons against LPS-induced endotoxemia may result from the apoE moiety and that human recombinant apoE may serve as a therapeuticum to protect against endotoxemia. PMID:9153287

  3. “Sticky” and “Promiscuous”—the Yin and Yang of Apolipoprotein A-I Termini in Discoidal High Density Lipoproteins: A Combined Computational-Experimental Approach†

    PubMed Central

    Jones, Martin K.; Gu, Feifei; Catte, Andrea; Li, Ling; Segrest, Jere P.

    2011-01-01

    Apolipoprotein (apo) A-I-containing lipoproteins in the form of high density lipoproteins (HDL) are inversely correlated with atherosclerosis. Because HDL is a soft form of condensed matter easily deformable by thermal fluctuations, the molecular mechanisms for HDL remodeling are not well understood. A promising approach to understanding HDL structure and dynamics is molecular dynamics (MD). In the present study, two computational strategies, MD simulated annealing (MDSA) and MD temperature-jump, were combined with experimental particle reconstitution to explore molecular mechanisms for phospholipid (PL)-rich HDL particle remodeling. The N-terminal domains of full length apoA-I were shown to be “sticky”, acting as a molecular latch largely driven by salt bridges, until, at a critical threshold of particle size, the associated domains released to expose extensive hydrocarbon regions of the PL to solvent. The “sticky” N-termini also associate with other apoA-I domains, perhaps being involved in N-terminal loops suggested by other laboratories. Alternatively, the overlapping helix 10 C-terminal domains of apoA-I were observed to be extremely mobile or “promiscuous”, transiently exposing limited hydrocarbon regions of PL. Based upon these models and reconstitution studies, we propose that separation of the N-terminal domains, as particles exceed a critical size, trigger fusion between particles or between particles and membranes, while the C-terminal domains of apoA-I drive the exchange of polar lipids down concentration gradients between particles. This hypothesis has significant biological relevance since lipid exchange and particle remodeling are critically important processes during metabolism of HDL particles at every step in the anti-atherogenic process of reverse cholesterol transport. PMID:21329368

  4. Complement Factor H Binds to Human Serum Apolipoprotein E and Mediates Complement Regulation on High Density Lipoprotein Particles.

    PubMed

    Haapasalo, Karita; van Kessel, Kok; Nissilä, Eija; Metso, Jari; Johansson, Tiira; Miettinen, Sini; Varjosalo, Markku; Kirveskari, Juha; Kuusela, Pentti; Chroni, Angelika; Jauhiainen, Matti; van Strijp, Jos; Jokiranta, T Sakari

    2015-11-27

    The alternative pathway of complement is an important part of the innate immunity response against foreign particles invading the human body. To avoid damage to host cells, it needs to be efficiently down-regulated by plasma factor H (FH) as exemplified by various diseases caused by mutations in its domains 19-20 (FH19-20) and 5-7 (FH5-7). These regions are also the main interaction sites for microbial pathogens that bind host FH to evade complement attack. We previously showed that inhibition of FH binding by a recombinant FH5-7 construct impairs survival of FH binding pathogens in human blood. In this study we found that upon exposure to full blood, the addition of FH5-7 reduces survival of, surprisingly, also those microbes that are not able to bind FH. This effect was mediated by inhibition of complement regulation and subsequently enhanced neutrophil phagocytosis by FH5-7. We found that although FH5-7 does not reduce complement regulation in the actual fluid phase of plasma, it reduces regulation on HDL particles in plasma. Using affinity chromatography and mass spectrometry we revealed that FH interacts with serum apolipoprotein E (apoE) via FH5-7 domains. Furthermore, binding of FH5-7 to HDL was dependent on the concentration of apoE on the HDL particles. These findings explain why the addition of FH5-7 to plasma leads to excessive complement activation and phagocytosis of microbes in full anticoagulated blood. In conclusion, our data show how FH interacts with apoE molecules via domains 5-7 and regulates alternative pathway activation on plasma HDL particles.

  5. Modification by acrolein, a component of tobacco smoke and age-related oxidative stress, mediates functional impairment of human apolipoprotein E.

    PubMed

    Tamamizu-Kato, Shiori; Wong, Jason Yiu; Jairam, Vikram; Uchida, Koji; Raussens, Vincent; Kato, Hiroyuki; Ruysschaert, Jean-Marie; Narayanaswami, Vasanthy

    2007-07-17

    Oxidative damage to proteins such as apolipoprotein B-100 increases the atherogenicity of low-density lipoproteins (LDL). However, little is known about the potential oxidative damage to apolipoprotein E (apoE), an exchangeable antiatherogenic apolipoprotein. ApoE plays an integral role in lipoprotein metabolism by regulating the plasma cholesterol and triglyceride levels. Hepatic uptake of lipoproteins is facilitated by apoE's ability to bind with cell surface heparan sulfate proteoglycans and to lipoprotein receptors via basic residues in its 22 kDa N-terminal domain (NT). We investigated the effect of acrolein, an aldehydic product of endogenous lipid peroxidation and a tobacco smoke component, on the conformation and function of recombinant human apoE3-NT. Acrolein caused oxidative modification of apoE3-NT as detected by Western blot with acrolein-lysine-specific antibodies, and tertiary conformational alterations. Acrolein modification impairs the ability of apoE3-NT to interact with heparin and the LDL receptor. Furthermore, acrolein-modified apoE3-NT displayed a 5-fold decrease in its ability to interact with lipid surfaces. Our data indicate that acrolein disrupts the functional integrity of apoE3, which likely interferes with its role in regulating plasma cholesterol homeostasis. These observations have implications regarding the role of apoE in the pathogenesis of smoking- and oxidative stress-mediated cardiovascular and cerebrovascular diseases.

  6. Calmodulin-like skin protein is downregulated in human cerebrospinal fluids of Alzheimer's disease patients with apolipoprotein E4; a pilot study using postmortem samples.

    PubMed

    Hashimoto, Yuichi; Umahara, Takahiko; Hanyu, Haruo; Iwamoto, Toshihiko; Matsuoka, Masaaki

    2017-09-01

    Calmodulin-like skin protein (CLSP) is a secreted peptide that inhibits neuronal cell death, linked to Alzheimer's disease (AD), by binding to the heterotrimeric humanin receptor and activating an intracellular survival pathway. CLSP is only expressed in skin keratinocytes and related epithelial cells, circulates in the blood stream, and passes the blood-cerebrospinal fluid (CSF) barrier. In the current study, we addressed the issues as to whether CLSP functions in the central nervous system and whether the concentration of CLSP is reduced in the CSFs of AD patients. Mice were intraperitoneally injected with 5 nmol of recombinant human CLSP. At 1h after the injection, the mice were sacrificed for the analysis of the existence of human CLSP in blood and interstitial fluid (ISF)-containing brain samples. Using postmortem CSF samples, we next determined the concentrations of CLSP in CSFs of human AD and control cases. Intraperitoneally administered recombinant human CLSP circulated in the blood stream and reached the brain interstitial fluid. The concentrations of CLSP in CSFs of human AD and control cases are sufficient to exhibit the CLSP activity. Although the concentrations of CLSP in CSFs were not significantly different between AD and control cases, the concentrations of CLSP are lower in the AD cases with the apolipoprotein E4 genotype than in the AD cases without the apolipoprotein E4 genotype. The first result indicates that CLSP enters the central nervous system through the blood-brain barrier. The second result suggests that CLSP functions in the human brains. The third result may exclude the possibility that the downregulation of the CLSP level is involved in the AD pathogenesis. The last result may contribute to the better understanding of the AD pathogenesis from the standpoint of the apolipoprotein E genotype.

  7. Structural Variation in Human Apolipoprotein E3 and E4: Secondary Structure, Tertiary Structure, and Size Distribution

    PubMed Central

    Chou, Chi-Yuan; Lin, Yi-Ling; Huang, Yu-Chyi; Sheu, Sheh-Yi; Lin, Ta-Hsien; Tsay, Huey-Jen; Chang, Gu-Gang; Shiao, Ming-Shi

    2005-01-01

    Human apolipoprotein E (apoE) is a 299-amino-acid protein with a molecular weight of 34 kDa. The difference between the apoE3 and apoE4 isoforms is a single residue substitution involving a Cys-Arg replacement at residue 112. ApoE4 is positively associated with atherosclerosis and late-onset and sporadic Alzheimer's disease (AD). ApoE4 and its C-terminal truncated fragments have been found in the senile plaques and neurofibrillary tangles in the brain of AD patients. However, detail structural information regarding isoform and domain interaction remains poorly understood. We prepared full-length, N-, and C-terminal truncated apoE3 and apoE4 proteins and studied their structural variation. Sedimentation velocity and continuous size distribution analysis using analytical ultracentrifugation revealed apoE372-299 as consisting of a major species with a sedimentation coefficient of 5.9. ApoE472-299 showed a wider and more complicated species distribution. Both apoE3 and E4 N-terminal domain (1–191) existed with monomers as the major component together with some tetramer. The oligomerization and aggregation of apoE protein increased when the C-terminal domain (192–271) was incorporated. The structural influence of the C-terminal domain on apoE is to assist self-association with no significant isoform preference. Circular dichroism and fluorescence studies demonstrated that apoE472-299 possessed a more α-helical structure with more hydrophobic residue exposure. The structural variation of the N-terminal truncated apoE3 and apoE4 protein provides useful information that helps to explain the greater aggregation of the apoE4 isoform and thus has implication for the involvement of apoE4 in AD. PMID:15475580

  8. An efficient on-column expressed protein ligation strategy: Application to segmental triple labeling of human apolipoprotein E3

    PubMed Central

    Zhao, Wentao; Zhang, Yonghong; Cui, Chunxian; Li, Qianqian; Wang, Jianjun

    2008-01-01

    Expressed protein ligation (EPL) is an intein-based approach that has been used for protein engineering and biophysical studies of protein structures. One major problem of the EPL is the low yield of final ligation product, primarily due to the complex procedure of the EPL, preventing EPL from gaining popularity in the research community. Here we report an efficient on-column EPL strategy, which focuses on enhancing the expression level of the intein-fusion protein that generates thioester for the EPL. We applied this EPL strategy to human apolipoprotein E (apoE) and routinely obtained 25–30 mg segmental, triple-labeled apoE from 1-L cell culture. The approaches reported here are general approaches that are not specific for apoE, thus providing a general strategy for a highly efficient EPL. In addition, we also report an isotopic labeling scheme that double-labels one domain and keeps the other domain of apoE deuterated. Such an isotopic labeling scheme can only be achieved using the EPL strategy. Our data indicated that the segmental triple-labeled apoEs using this labeling scheme produced high-quality, simplified NMR spectra, facilitating NMR spectral assignment. For large proteins, such as apoE, perdeuterated protein samples have to be used to reduce the linewidth of NMR signals, causing a major problem for the NOE-based NMR method, since perdeuterated proteins lack protons for NOE measurement. The new labeling strategy solves this problem and provides 13C/15N double-labeled, protonated protein domains, allowing for determination of high-resolution NMR structure of these large proteins. PMID:18305193

  9. Association of High-Density Lipoprotein-Cholesterol Versus Apolipoprotein A-I With Risk of Coronary Heart Disease: The European Prospective Investigation Into Cancer-Norfolk Prospective Population Study, the Atherosclerosis Risk in Communities Study, and the Women's Health Study.

    PubMed

    van Capelleveen, Julian C; Bochem, Andrea E; Boekholdt, S Matthijs; Mora, Samia; Hoogeveen, Ron C; Ballantyne, Christie M; Ridker, Paul M; Sun, Wensheng; Barter, Philip J; Tall, Alan R; Zwinderman, Aeilko H; Kastelein, John J P; Wareham, Nick J; Khaw, Kay-Tee; Hovingh, G Kees

    2017-08-03

    The contribution of apolipoprotein A-I (apoA-I) to coronary heart disease (CHD) risk stratification over and above high-density lipoprotein cholesterol (HDL-C) is unclear. We studied the associations between plasma levels of HDL-C and apoA-I, either alone or combined, with risk of CHD events and cardiovascular risk factors among apparently healthy men and women. HDL-C and apoA-I levels were measured among 17 661 participants of the EPIC (European Prospective Investigation into Cancer)-Norfolk prospective population study. Hazard ratios for CHD events and distributions of risk factors were calculated by quartiles of HDL-C and apoA-I. Results were validated using data from the ARIC (Atherosclerosis Risk in Communities) and WHS (Women's Health Study) cohorts, comprising 15 494 and 27 552 individuals, respectively. In EPIC-Norfolk, both HDL-C and apoA-I quartiles were strongly and inversely associated with CHD risk. Within HDL-C quartiles, higher apoA-I levels were not associated with lower CHD risk; in fact, CHD risk was higher within some HDL-C quartiles. ApoA-I levels were associated with higher levels of CHD risk factors: higher body mass index, HbA1c, non-HDL-C, triglycerides, apolipoprotein B, systolic blood pressure, and C-reactive protein, within fixed HDL-C quartiles. In contrast, HDL-C levels were consistently inversely associated with overall CHD risk and CHD risk factors within apoA-I quartiles (P<0.001). These findings were validated in the ARIC and WHS cohorts. Our findings demonstrate that apoA-I levels do not offer predictive information over and above HDL-C. In fact, within some HDL-C quartiles, higher apoA-I levels were associated with higher risk of CHD events, possibly because of the unexpected higher prevalence of cardiovascular risk factors in association with higher apoA-I levels. URL: https://www.clinicaltrials.gov. Unique identifier: NCT00000479. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

  10. [Genetic polymorphism of the E apolipoprotein in school age children: comparison with levels of plasma lipids and apolipoproteins].

    PubMed

    Callas, Ney; Poveda, Elpidia; Baracaldo, César; Hernández, Patricia; Castillo, Carlina; Guerra, Martha

    2007-12-01

    Research in laboratories around the world has documented the contribution of the E apolipoprotein alleles to structural variations of lipids and apolipoproteins. The gene frequencies of the E apolipoprotein alleles were compared with the lipid and apolipoprotein levels in school age children. Six hundred and ninety one 5 to 15 years old school age children from the Colombian departments of Cundinamarca, Boyacá, Meta, Santander and Norte de Santander, were evaluated. The genotypes of the E apolipoprotein were identified by polymerase chain reaction-restriction fragment length polymorphism. Plasma levels for the following 5 lipids and lipoproteins were assayed: total cholesterol, HDL (high density lipoprotein) cholesterol, LDL (low density lipoprotein) cholesterol, triglycerides, VLDL (very low density lipoprotein) cholesterol, A-I apolipoprotein and B-100 apolipoprotein. Alleles e2, e3 and e4 were found in frequencies of 0.04, 0.86 and 0.08, respectively. The E4 group (E4/3-E4/4), in contrast with the E2 group (E3/2-E2/2), presented highest plasma concentrations of total cholesterol, LDL cholesterol and B-100 apolipoprotein (p=0.014, 0.001 and 0.000, respectively). When the E3/3 group was compared with E2, the same result was obtained (p=0.015, 0.002 and 0.002, respectively). The influence of the E apolipoprotein polymorphism appeared greater in female children than male. The e4 allele was associated with higher levels of total cholesterol, LDL cholesterol and B-100 apolipoprotein and indicates the necessity of additional research into the interactions between polymorphism E apolipoprotein and other genes, life styles, risk factors and potential contribution to cardiovascular diseases.

  11. Role of apolipoprotein E in febrile convulsion.

    PubMed

    Giray, Ozlem; Ulgenalp, Ayfer; Bora, Elçin; Uran, Nedret; Yilmaz, Ebru; Unalp, Aycan; Erçal, Derya

    2008-10-01

    Apolipoprotein E is consistently associated with the progression of some common human neurodegenerative diseases, e.g., epilepsy. We hypothesized that genetic variations in the apolipoprotein E gene have implications for susceptibility to, and prognoses in, febrile convulsion, which plays an apparent role in the development of epilepsy. We used the polymerase chain reaction and restriction enzyme digestion to characterize variations of the apolipoprotein E gene. Sixty-nine patients with febrile convulsion (simple/complex) and a corresponding cohort of healthy patients (n = 75) were used. There was no significant difference in genotypic distribution and allelic frequencies of the apolipoprotein E gene between the febrile convulsion and control groups. Comparing subpopulations of the febrile convulsion group (patients with simple and complex febrile convulsion), we noted that no patients with the epsilon3/epsilon4 genotype had complex febrile convulsions. The apolipoprotein E epsilon3/epsilon4 genotype was more frequently seen in the simple febrile than in the complicated febrile convulsion group (9 versus 0 patients, respectively). The data indicate an association with the epsilon3/epsilon4 genotype of the apolipoprotein E gene with a milder phenotype. Although apolipoprotein E4 is not a vulnerability factor regarding febrile convulsions, it seems effective in regard to prognoses.

  12. Real time magnetic resonance imaging of apo AI metabolism in vivo

    USDA-ARS?s Scientific Manuscript database

    Apolipoprotein AI (apo AI) plays a key role in maintaining cardiovascular health and constitutes the major lipoprotein component in high density lipoproteins (HDL). It metabolism, however, follows a complex pathway. Synthesized in the liver and intestines, its becomes lipidated by its interaction w...

  13. Human factors involvement in bringing the power of AI to a heterogeneous user population

    NASA Technical Reports Server (NTRS)

    Czerwinski, Mary; Nguyen, Trung

    1994-01-01

    The Human Factors involvement in developing COMPAQ QuickSolve, an electronic problem-solving and information system for Compaq's line of networked printers, is described. Empowering customers with expert system technology so they could solve advanced networked printer problems on their own was a major goal in designing this system. This process would minimize customer down-time, reduce the number of phone calls to the Compaq Customer Support Center, improve customer satisfaction, and, most importantly, differentiate Compaq printers in the marketplace by providing the best, and most technologically advanced, customer support. This represents a re-engineering of Compaq's customer support strategy and implementation. In its first generation system, SMART, the objective was to provide expert knowledge to Compaq's help desk operation to more quickly and correctly answer customer questions and problems. QuickSolve is a second generation system in that customer support is put directly in the hands of the consumers. As a result, the design of QuickSolve presented a number of challenging issues. Because the produce would be used by a diverse and heterogeneous set of users, a significant amount of human factors research and analysis was required while designing and implementing the system. Research that shaped the organization and design of the expert system component as well.

  14. Antisense inhibition of apolipoprotein (a) to lower plasma lipoprotein (a) levels in humans

    PubMed Central

    Graham, Mark J.; Viney, Nick; Crooke, Rosanne M.; Tsimikas, Sotirios

    2016-01-01

    Epidemiological, genetic association, and Mendelian randomization studies have provided strong evidence that lipoprotein (a) [Lp(a)] is an independent causal risk factor for CVD, including myocardial infarction, stroke, peripheral arterial disease, and calcific aortic valve stenosis. Lp(a) levels >50 mg/dl are highly prevalent (20% of the general population) and are overrepresented in patients with CVD and aortic stenosis. These data support the notion that Lp(a) should be a target of therapy for CVD event reduction and to reduce progression of aortic stenosis. However, effective therapies to specifically reduce plasma Lp(a) levels are lacking. Recent animal and human studies have shown that Lp(a) can be specifically targeted with second generation antisense oligonucleotides (ASOs) that inhibit apo(a) mRNA translation. In apo(a) transgenic mice, an apo(a) ASO reduced plasma apo(a)/Lp(a) levels and their associated oxidized phospholipid (OxPL) levels by 86 and 93%, respectively. In cynomolgus monkeys, a second generation apo(a) ASO, ISIS-APO(a)Rx, significantly reduced hepatic apo(a) mRNA expression and plasma Lp(a) levels by >80%. Finally, in a phase I study in normal volunteers, ISIS-APO(a)Rx ASO reduced Lp(a) levels and their associated OxPL levels up to 89 and 93%, respectively, with minimal effects on other lipoproteins. ISIS-APO(a)Rx represents the first specific and potent drug in clinical development to lower Lp(a) levels and may be beneficial in reducing CVD events and progression of calcific aortic valve stenosis. PMID:26538546

  15. Identification and characterization of a new human gene (APOC4) in the apolipoprotein E, C-I, and C-II gene locus

    SciTech Connect

    Allan, C.M.; Walker, D.; Taylor, J.M.; Segrest, J.P.

    1995-07-20

    We have identified and characterized a previously unreported human gene that is found within the apolipoprotein (apo) E/C-I/C-II gene locus. On the basis of its location and its properties, this new gene has been designated APOC4. Nucleotide sequence analysis of genomic DNA and liver cDNA clones revealed a 3.3-kb gene consisting of three exons and two introns. Its 3{prime} terminus lies 555 bp upstream of APOC2, giving both genes the same transcriptional orientation. The promoter of the APOC4 gene lacks a typical TATA box, consistent with an apparent heterogeneity in transcription start sites. RNase protection analysis indicated relatively low apoC-IV mRNA levels in human liver, compared to apoC-II mRNA levels. The predicted apoC-IV protein sequence, comprising 127 amino acid residues, contains a putative 25-residue signal peptide and two potential amphipathic {alpha}-helical domains. Amino acid sequence comparisons indicate a limited homology between apoC-IV and either apoC-I or apoC-II. Since its hepatic expression and predicted protein structure are characteristic of the other genes in this cluster, we propose that the APOC4 gene is a member of the apolipoprotein gene family. 53 refs., 6 figs.

  16. Separation and isolation of human apolipoproteins C-II, C-III0, C-III1, and C-III2 by chromatofocusing on the Fast Protein Liquid Chromatography system.

    PubMed

    Huff, M W; Strong, W L

    1987-09-01

    Chromatofocusing, which separates proteins based on differences in isoelectric point, has been used on the Fast Protein Liquid Chromatography (FPLC) system (Pharmacia) to separate the C apolipoproteins from human very low density lipoproteins (VLDL). Using a Mono P column (Pharmacia), a pH gradient between pH 6.2 and pH 4.0 was generated using buffers containing 6 M urea, at a flow rate of 0.5 ml/min. Typically, runs took approximately 45 min. Chromatofocusing of delipidated whole VLDL produced sharp, well-resolved peaks for the C apolipoproteins. However, as determined by analytical isoelectric focusing (IEF), the apolipoprotein E isoforms were not separated from apoC-II, and they contaminated the other apoC species to a variable extent. In addition, apoC-II was not resolved from apoC-III0. Preliminary precipitation of VLDL with acetone prior to delipidation removed both apolipoproteins E and B. Using a start buffer of 25 mM histidine, pH 6.2, and a 1:30 dilution of the polybuffer exchanger (eluting buffer), apoC-II, C-III0, C-III1, and C-III2 were well resolved in run-times of approximately 60 min. The C apoproteins proved to be pure by analytical IEF and immunoassay with monospecific antisera against apoC-II and C-III. Recovery was over 90% of the protein chromatographed. In addition, a variant of apoC-II present in VLDL of a hypertriglyceridemic subject was clearly resolved from the other C apolipoproteins. This technique is superior to conventional methodology in terms of its time saving and high resolution. The application of this technique to the study of C apolipoprotein variants and C apolipoprotein specific radioactivity determinations is possible.

  17. A pharmacogenomic profile of human neural progenitors undergoing differentiation in the presence of the traditional Chinese medicine NeuroAiD.

    PubMed

    Chan, H Y A; Stanton, L W

    2016-10-01

    NeuroAiD, a traditional Chinese medicine widely used to treat stroke patients in China, was recently demonstrated in rodent models and in clinical trials to possess neuroregenerative and neuroprotective properties. In order to understand the mechanisms employed by NeuroAiD to bring about its neuroproliferative and neuroprotective effects, we investigated the impact of MLC901, a reformulated version of MLC601, on human neural progenitors undergoing neural differentiation at the molecular level by performing three independent microarray experiments. Functional annotations of the genes regulated by MLC901 that were associated with neurogenesis were found to be enriched. We also identified potential targets (FGF19, GALR2, MMP10, FGF3 and TDO2) of MLC901 that could promote neurogenesis and neuroprotection in the human brain. This work highlighted some interesting targets and offered some insights into the possible mechanism of action of MLC901. The discovery could also provide a platform to the development of future therapeutic targets.

  18. Fast protein chromatofocusing of human very-low-density lipoproteins.

    PubMed

    Weisweiler, P; Friedl, C; Schwandt, P

    1986-01-03

    Using fast protein chromatofocusing, a high-efficiency column chromatography method with a self-generated pH gradient and focusing effects, soluble human very-low-density lipoprotein (VLDL) apolipoproteins were fractionated between pH 6.3 and 4.0. In the presence of 6 mol/l urea and with a flow rate of 1 ml/min, one run (up to 10 mg of protein) took 30 min. VLDL apolipoproteins were separated in seven peaks. As revealed by SDS-polyacrylamide gel electrophoresis, isoelectric focusing and double-immunodiffusion against mono-specific antisera, fractions corresponded to the following proteins: apolipoprotein C-I, albumin, apolipoproteins A-I, E, C-II plus C-III0, C-III1 and C-III2, respectively. Apolipoproteins were eluted in sharp, well-resolved peaks. The recovery of proteins was 78% of the starting material. With fast protein chromatofocusing, an efficient isolation of single apolipoproteins is possible from small amounts of VLDL apolipoprotein preparations. This technique is superior to the commonly used, time-consuming methods for apolipoprotein isolation.

  19. Dietary cholesterol and its effect on tau protein: a study in apolipoprotein E-deficient and P301L human tau mice.

    PubMed

    Glöckner, Frauke; Meske, Volker; Lütjohann, Dieter; Ohm, Thomas G

    2011-04-01

    Apolipoprotein E (ApoE) is the major cholesterol transporter in the brain. There is epidemiological and experimental evidence for involvement of cholesterol metabolism in the development and progression of Alzheimer disease. A dietary effect on tau phosphorylation or aggregation, or a role of apoE in tau metabolism, has been studied experimentally, but the data are ambiguous. To elucidate the relationship between cholesterol and tau, we studied mice expressing P301L mutant human tau but not apoE (htau-ApoE) and P301L mice with wild-type ApoE (htau- ApoE); both genotypes develop neuron cytoskeletal changes similar to those found in Alzheimer disease. Mice were kept on a cholesterol-enriched diet or control diet for 15 weeks. The numbers of neurons with hyperphosphorylated and conformationally changed tau in the cerebral cortex were assessed by immunohistochemistry, and sterol levels were determined. Highly elevated dietary serum cholesterol levels enhanced ongoing tau pathology in htau-ApoE mice; this effect correlated with elevated brain cholesterol metabolite 27-hydroxycholesterol levels. Apolipoprotein E deficiency promoted significant increases of tau phosphorylation and conformational changes in mice on a control diet. In htau-ApoE mice on the high cholesterol regimen, brain oxysterol levels were less than in htau-ApoE mice, and the numbers of neurons with pathologically altered tau were similar to those in htau-ApoE mice on the high-cholesterol diet.

  20. Analytical isoelectric focusing of apolipoprotein B of human plasma low-density lipoproteins in the presence of a nonionic and a zwitterionic detergent.

    PubMed

    Melnik, B C; Melnik, S F

    1988-06-01

    A method for the analytical isoelectric focusing of Nonidet-P40-delipidated apolipoprotein B of human plasma low-density lipoproteins has been developed. Isoelectric focusing was performed in the presence of the zwitterionic nondenaturing detergent Chaps, 3-[(3-cholamidopropyl)-dimethylammonio]-1-propane sulfonate, and the nonionic surfactant Nonidet-P40, polyoxyethyleneglycol p-t-octylphenol with a mean of 9.0 ethylene oxide units per molecule. Low-density lipoprotein (LDL) apolipoprotein B (apo-B) entered 3.75% polyacrylamide gels without precipitation at the sites of sample application, permitting apoprotein recoveries of greater than 90% in the migrating bands. LDL apo-B exhibited 10 distinguishable bands with apparent isoelectric points of 7.34 (band 1), 7.27 (band 2), 7.16 (band 3), 7.02 (band 4), 6.88 (band 5), 6.70 (band 6), 6.61 (band 7), 6.48 (band 8), 6.40 (band 9), and 6.24 (band 10), respectively. Bands 3 and 4, 6 and 7, as well as 8 and 9 could be identified as major double bands. When the focused apo-B was run in a second dimension by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the same relative molecular weight of B-100 was obtained for all focused bands. After electrotransfer to nitrocellulose paper, all bands reacted with polyclonal anti-human LDL antibody. Furthermore, the detergent-solubilized apo-B retained the immunological properties of native low-density lipoproteins when tested by double immunodiffusion against polyvalent anti-human LDL sera.

  1. Apolipoprotein B100

    MedlinePlus

    ... skin is broken) Multiple punctures to locate veins Considerations Apolipoprotein measurements may provide more detail about your risk for heart disease, but the added value of this test beyond a lipid panel is ...

  2. Targeting nanodisks via a single chain variable antibody - Apolipoprotein chimera

    SciTech Connect

    Iovannisci, David M.; Beckstead, Jennifer A.; Ryan, Robert O.

    2009-02-06

    Nanodisks (ND) are nanometer scale complexes of phospholipid and apolipoprotein that have been shown to function as drug delivery vehicles. ND harboring significant quantities of the antifungal agent, amphotericin B, or the bioactive isoprenoid, all trans retinoic acid, have been generated and characterized. As currently formulated, ND possess limited targeting capability. In this study, we constructed a single chain variable antibody (scFv).apolipoprotein chimera and assessed the ability of this fusion protein to form ND and recognize the antigen to which the scFv is directed. Data obtained revealed that {alpha}-vimentin scFv.apolipoprotein A-I is functional in ND formation and antigen recognition, opening the door to the use of such chimeras in targeting drug-enriched ND to specific tissues.

  3. Targeting nanodisks via a single chain variable antibody -apolipoprotein chimera*

    PubMed Central

    Iovannisci, David M.; Beckstead, Jennifer A.; Ryan, Robert O.

    2009-01-01

    Nanodisks (ND) are nanometer scale complexes of phospholipid and apolipoprotein that have been shown to function as drug delivery vehicles. ND harboring significant quantities of the antifungal agent, amphotericin B, or the bioactive isoprenoid, all trans retinoic acid, have been generated and characterized. As currently formulated, ND possess limited targeting capability. In the present study we constructed a single chain variable antibody (scFv)•apolipoprotein chimera and assessed the ability of this fusion protein to form ND and recognize the antigen to which the scFv is directed. Data obtained revealed that α-vimentin scFv•apolipoprotein A-I is functional in ND formation and antigen recognition, opening the door to the use of such chimeras in targeting drug-enriched ND to specific tissues. PMID:19114030

  4. Conformational studies of the N-terminal lipid-associating domain of human apolipoprotein C-I by CD and 1H NMR spectroscopy.

    PubMed Central

    Rozek, A.; Buchko, G. W.; Kanda, P.; Cushley, R. J.

    1997-01-01

    A peptide comprising the N-terminal 38 residues of human apolipoprotein C-I (apoC-I(1-38)) was synthesized using solid-phase methods and its solution conformation studied by CD and 1H NMR spectroscopy. The CD data indicate that apoC-I(1-38) has a similar helical content (55%) in the presence of saturating amounts of SDS or egg yolk lysophosphatidylcholine. A structural ensemble of SDS-bound apoC-I(1-38) was calculated from 464 NOE-based distance restraints using distance geometry methods. ApoC-I(1-38) adopts a helical structure between residues V4 and K30 and an extended C-terminus from Q31 when associated with SDS. The region K12-G15 undergoes slow conformational exchange as indicated by above-average amide resonance linewidths, large temperature coefficients, and fast exchange (< 2 h) of backbone amide protons with deuterium. The mobility of K12-G15 is reflected in the poorly defined dihedral angles of K12 and E13 in the calculated ensemble of structures. The average structure of apoC-I(1-38) is curved toward its hydrophobic face with bends of 125 degrees, centered at K12/E13, and 150 degrees, centered at K21. This curvature appears to be driven by the interaction of two hydrophobic clusters, one formed by residues L8, L11, F14, and L18, and the other by L25, I26, and I29, with the amphiphile SDS. Based on our present structural definition of apoC-I(1-38) and the previously obtained structure of the fragment apoC-I(35-53), we propose the secondary structure of intact apolipoprotein C-I. PMID:9300485

  5. Plasma Lipoproteins as Mediators of the Oxidative Stress Induced by UV Light in Human Skin: A Review of Biochemical and Biophysical Studies on Mechanisms of Apolipoprotein Alteration, Lipid Peroxidation, and Associated Skin Cell Responses

    PubMed Central

    Filipe, Paulo; Morlière, Patrice; Silva, João N.; Mazière, Jean-Claude; Patterson, Larry K.; Freitas, João P.; Santus, R.

    2013-01-01

    There are numerous studies concerning the effect of UVB light on skin cells but fewer on other skin components such as the interstitial fluid. This review highlights high-density lipoprotein (HDL) and low-density lipoprotein (LDL) as important targets of UVB in interstitial fluid. Tryptophan residues are the sole apolipoprotein residues absorbing solar UVB. The UVB-induced one-electron oxidation of Trp produces •Trp and •O2 − radicals which trigger lipid peroxidation. Immunoblots from buffered solutions or suction blister fluid reveal that propagation of photooxidative damage to other residues such as Tyr or disulfide bonds produces intra- and intermolecular bonds in apolipoproteins A-I, A-II, and B100. Partial repair of phenoxyl tyrosyl radicals (TyrO•) by α-tocopherol is observed with LDL and HDL on millisecond or second time scales, whereas limited repair of α-tocopherol by carotenoids occurs in only HDL. More effective repair of Tyr and α-tocopherol is observed with the flavonoid, quercetin, bound to serum albumin, but quercetin is less potent than new synthetic polyphenols in inhibiting LDL lipid peroxidation or restoring α-tocopherol. The systemic consequences of HDL and LDL oxidation and the activation and/or inhibition of signalling pathways by oxidized LDL and their ability to enhance transcription factor DNA binding activity are also reviewed. PMID:23738035

  6. Baculovirus-mediated expression of human apolipoprotein E in Manduca sexta larvae generates particles that bind to the low density lipoprotein receptor.

    PubMed Central

    Gretch, D G; Sturley, S L; Friesen, P D; Beckage, N E; Attie, A D

    1991-01-01

    Human apolipoprotein E (apoE) is a ligand for the low density lipoprotein (LDL) receptor and mediates the catabolism of several classes of lipoprotein particles. Binding of apoE to the LDL receptor requires association of apoE with lipid in a vesicle or a lipoprotein particle. Because of this requirement, purified apoE or apoE derived directly from bacterial expression systems does not bind to the LDL receptor. To overcome this problem and to facilitate analysis of apoE structure, recombinant baculoviruses containing the human apoE cDNA fused to the polyhedrin promoter of Autographa californica nuclear polyhedrosis virus were constructed. The recombinant viruses were used to infect larvae of the tobacco hornworm Manduca sexta in vivo. High levels of lipoprotein particles containing human apoE were present in the hemolymph of infected larvae. In contrast to apoE produced by recombinant baculovirus-infected insect cells in vitro, these particles were excellent ligands for the LDL receptor. Images PMID:1924311

  7. Clinical significance of apolipoprotein A5

    USDA-ARS?s Scientific Manuscript database

    We have investigated the evidence from recent human studies examining the role of apolipoprotein A-V (APOA-V) in triglyceride-rich lipoprotein metabolism and cardiovascular disease (CVD) risk. Special emphasis was placed on the evidence emerging from the association between genetic variability at th...

  8. Characterization of five new mutants in the carboxyl-terminal domain of human apolipoprotein E: No cosegregation with severe hyperlipidemia

    SciTech Connect

    Maagdenberg, A.M.J.M. van den; Bruijn, I.H. de; Hofker, M.H.; Frants, R.R. ); Knijff, P. de; Smelt, A.H.M.; Leuven, J.A.G.; van't Hooft, F.; Assmann, G.; Havekes, L.M. ); Weng, Wei; Funke, H. )

    1993-05-01

    Assessment of the apolipoprotein E (apoE) phenotype by isoelectric focusing of both hyperlipidemic and normolipidemic individuals identified five new variants. All mutations were confined to the downstream part of the APOE gene by using denaturing gradient gel electrophoresis (DGGE). Sequence analysis revealed five new mutations causing unique amino acid substitutions in the carboxyl-terminal part of the protein containing the putative lipid-binding domain. Three hyperlipoproteinemic probands were carriers of the APOE*2(Va1236[r arrow]Glu) allele, the APOE*3(Cys112-Arg; Arg251[r arrow]Gly) allele, or the APOE*1(Arg158[r arrow]Cys; Leu252[r arrow]Glu) allele. DGGE of the region encoding the receptor-binding domain was useful for haplotyping the mutations at codons 112 and 158. Family studies failed to demonstrate cosegregation between the new mutations and severe hyperlipoproteinemia, although a number of carriers for the APOE*3(Cys112[r arrow]Arg; Arg251[r arrow]Gly) allele and the APOE*1(Arg158-Cys; Leu252[r arrow]Glu) allele expressed hypertriglyceridemia and/ or hypercholesterolemia. Two other mutant alleles, APOE*4[sup [minus

  9. Artificial Intelligence Study (AIS).

    DTIC Science & Technology

    1987-02-01

    ARTIFICIAL INTELLIGNECE HARDWARE ....... 2-50 AI Architecture ................................... 2-49 AI Hardware ....................................... 2...system: The synergy between discrete-event simulation and the approaches that programmers take to develop artifical - intelligence software took some...dated 28 October 1986 Subject: Army Artifical Intelligence Training Available at Fort Gordon) has indicated the availability of the following training

  10. Sesamin attenuates intercellular cell adhesion molecule-1 expression in vitro in TNF-alpha-treated human aortic endothelial cells and in vivo in apolipoprotein-E-deficient mice.

    PubMed

    Wu, Wen-Huey; Wang, Shu-Huei; Kuan, I-I; Kao, Ya-Shi; Wu, Pei-Jhen; Liang, Chan-Jung; Chien, Hsiung-Fei; Kao, Chiu-Hua; Huang, Ching-Jang; Chen, Yuh-Lien

    2010-09-01

    Sesame lignans have antioxidative and anti-inflammatory properties. We focused on the effects of the lignans sesamin and sesamol on the expression of endothelial-leukocyte adhesion molecules in tumor necrosis factor-alpha (TNF-alpha)-treated human aortic endothelial cells (HAECs). When HAECs were pretreated with sesamin (10 or 100 microM), the TNF-alpha-induced expression of intercellular cell adhesion molecule-1 (ICAM-1) was significantly reduced (35 or 70% decrease, respectively) by Western blotting. Sesamol was less effective at inhibiting ICAM-1 expression (30% decrease at 100 microM). Sesamin and sesamol reduced the marked TNF-alpha-induced increase in human antigen R (HuR) translocation and the interaction between HuR and the 3'UTR of ICAM-1 mRNA. Both significantly reduced the binding of monocytes to TNF-alpha-stimulated HAECs. Sesamin significantly attenuated TNF-alpha-induced ICAM-1 expression and cell adhesion by downregulation of extracellular signal-regulated kinase 1/2 and p38. Furthermore, in vivo, sesamin attenuated intimal thickening and ICAM-1 expression seen in aortas of apolipoprotein-E-deficient mice. Taken together, these data suggest that sesamin inhibits TNF-alpha-induced extracellular signal-regulated kinase/p38 phosphorylation, nuclear translocation of NF-kappaB p65, cytoplasmic translocalization of HuR and thereby suppresses ICAM-1 expression, resulting in reduced adhesion of leukocytes. These results also suggest that sesamin may prevent the development of atherosclerosis and inflammatory responses.

  11. Sex-dependent modulation of longevity by two Drosophila homologues of human Apolipoprotein D, GLaz and NLaz.

    PubMed

    Ruiz, Mario; Sanchez, Diego; Canal, Inmaculada; Acebes, Angel; Ganfornina, Maria D

    2011-07-01

    Apolipoprotein D (ApoD), a member of the Lipocalin family, is the gene most up-regulated with age in the mammalian brain. Its expression strongly correlates with aging-associated neurodegenerative and metabolic diseases. Two homologues of ApoD expressed in the Drosophila brain, Glial Lazarillo (GLaz) and Neural Lazarillo (NLaz), are known to alter longevity in male flies. However, sex differences in the aging process have not been explored so far for these genes. Here we demonstrate that NLaz alters lifespan in both sexes, but unexpectedly the lack of GLaz influences longevity in a sex-specific way, reducing longevity in males but not in females. While NLaz has metabolic functions similar to ApoD, the regulation of GLaz expression upon aging is the closest to ApoD in the aging brain. A multivariate analysis of physiological parameters relevant to lifespan modulation uncovers both common and specialized functions for the two Lipocalins, and reveals that changes in protein homeostasis account for the observed sex-specific patterns of longevity. The response to oxidative stress and accumulation of lipid peroxides are among their common functions, while the transcriptional and behavioral response to starvation, the pattern of daily locomotor activity, storage of fat along aging, fertility, and courtship behavior differentiate NLaz from GLaz mutants. We also demonstrate that food composition is an important environmental parameter influencing stress resistance and reproductive phenotypes of both Lipocalin mutants. Since ApoD shares many properties with the common ancestor of invertebrate Lipocalins, we must benefit from this global comparison with both GLaz and NLaz to understand the complex functions of ApoD in mammalian aging and neurodegeneration. Copyright © 2011 Elsevier Inc. All rights reserved.

  12. Zebrafish as a model for apolipoprotein biology: comprehensive expression analysis and a role for ApoA-IV in regulating food intake

    PubMed Central

    Otis, Jessica P.; Zeituni, Erin M.; Thierer, James H.; Anderson, Jennifer L.; Brown, Alexandria C.; Boehm, Erica D.; Cerchione, Derek M.; Ceasrine, Alexis M.; Avraham-Davidi, Inbal; Tempelhof, Hanoch; Yaniv, Karina; Farber, Steven A.

    2015-01-01

    Improved understanding of lipoproteins, particles that transport lipids throughout the circulation, is vital to developing new treatments for the dyslipidemias associated with metabolic syndrome. Apolipoproteins are a key component of lipoproteins. Apolipoproteins are proteins that structure lipoproteins and regulate lipid metabolism through control of cellular lipid exchange. Constraints of cell culture and mouse models mean that there is a need for a complementary model that can replicate the complex in vivo milieu that regulates apolipoprotein and lipoprotein biology. Here, we further establish the utility of the genetically tractable and optically clear larval zebrafish as a model of apolipoprotein biology. Gene ancestry analyses were implemented to determine the closest human orthologs of the zebrafish apolipoprotein A-I (apoA-I), apoB, apoE and apoA-IV genes and therefore ensure that they have been correctly named. Their expression patterns throughout development were also analyzed, by whole-mount mRNA in situ hybridization (ISH). The ISH results emphasized the importance of apolipoproteins in transporting yolk and dietary lipids: mRNA expression of all apolipoproteins was observed in the yolk syncytial layer, and intestinal and liver expression was observed from 4–6 days post-fertilization (dpf). Furthermore, real-time PCR confirmed that transcription of three of the four zebrafish apoA-IV genes was increased 4 hours after the onset of a 1-hour high-fat feed. Therefore, we tested the hypothesis that zebrafish ApoA-IV performs a conserved role to that in rat in the regulation of food intake by transiently overexpressing ApoA-IVb.1 in transgenic larvae and quantifying ingestion of co-fed fluorescently labeled fatty acid during a high-fat meal as an indicator of food intake. Indeed, ApoA-IVb.1 overexpression decreased food intake by approximately one-third. This study comprehensively describes the expression and function of eleven zebrafish apolipoproteins

  13. Zebrafish as a model for apolipoprotein biology: comprehensive expression analysis and a role for ApoA-IV in regulating food intake.

    PubMed

    Otis, Jessica P; Zeituni, Erin M; Thierer, James H; Anderson, Jennifer L; Brown, Alexandria C; Boehm, Erica D; Cerchione, Derek M; Ceasrine, Alexis M; Avraham-Davidi, Inbal; Tempelhof, Hanoch; Yaniv, Karina; Farber, Steven A

    2015-03-01

    Improved understanding of lipoproteins, particles that transport lipids throughout the circulation, is vital to developing new treatments for the dyslipidemias associated with metabolic syndrome. Apolipoproteins are a key component of lipoproteins. Apolipoproteins are proteins that structure lipoproteins and regulate lipid metabolism through control of cellular lipid exchange. Constraints of cell culture and mouse models mean that there is a need for a complementary model that can replicate the complex in vivo milieu that regulates apolipoprotein and lipoprotein biology. Here, we further establish the utility of the genetically tractable and optically clear larval zebrafish as a model of apolipoprotein biology. Gene ancestry analyses were implemented to determine the closest human orthologs of the zebrafish apolipoprotein A-I (apoA-I), apoB, apoE and apoA-IV genes and therefore ensure that they have been correctly named. Their expression patterns throughout development were also analyzed, by whole-mount mRNA in situ hybridization (ISH). The ISH results emphasized the importance of apolipoproteins in transporting yolk and dietary lipids: mRNA expression of all apolipoproteins was observed in the yolk syncytial layer, and intestinal and liver expression was observed from 4-6 days post-fertilization (dpf). Furthermore, real-time PCR confirmed that transcription of three of the four zebrafish apoA-IV genes was increased 4 hours after the onset of a 1-hour high-fat feed. Therefore, we tested the hypothesis that zebrafish ApoA-IV performs a conserved role to that in rat in the regulation of food intake by transiently overexpressing ApoA-IVb.1 in transgenic larvae and quantifying ingestion of co-fed fluorescently labeled fatty acid during a high-fat meal as an indicator of food intake. Indeed, ApoA-IVb.1 overexpression decreased food intake by approximately one-third. This study comprehensively describes the expression and function of eleven zebrafish apolipoproteins and

  14. Cross-species pharmacokinetic comparison from mouse to man of a second-generation antisense oligonucleotide, ISIS 301012, targeting human apolipoprotein B-100.

    PubMed

    Yu, Rosie Z; Kim, Tae-Won; Hong, An; Watanabe, Tanya A; Gaus, Hans J; Geary, Richard S

    2007-03-01

    The pharmacokinetics of a 2'-O-(2-methoxyethyl)-modified oligonucleotide, ISIS 301012 [targeting human apolipoprotein B-100 (apoB-100)], was characterized in mouse, rat, monkey, and human. Plasma pharmacokinetics following parental administration was similar across species, exhibiting a rapid distribution phase with t(1/2alpha) of several hours and a prolonged elimination phase with t(1/2beta) of days. The prolonged elimination phase represents equilibrium between tissues and circulating drug due to slow elimination from tissues. Absorption was nearly complete following s.c. injection, with bioavailability ranging from 80 to 100% in monkeys. Plasma clearance scaled well across species as a function of body weight alone, and this correlation was improved when corrected for plasma protein binding. In all of the animal models studied, the highest tissue concentrations of ISIS 301012 were observed in kidney and liver. Urinary excretion was less than 3% in monkeys and human in the first 24 h. ISIS 301012 is highly bound to plasma proteins, probably preventing rapid removal by renal filtration. However, following 25 mg/kg s.c. administration in mouse and 5-mg/kg i.v. bolus administration in rat, plasma concentrations of ISIS 301012 exceeded their respective protein binding capacity. Thus, urinary excretion increased to 16% or greater within the first 24 h. Albeit slow, urinary excretion of ISIS 301012 and its shortened metabolites is the ultimate elimination pathway of this compound, as demonstrated by 32% of dose recovered in total excreta by 14 days in a rat mass balance study. The pharmacokinetics of ISIS 301012 in human is predictable from the pharmacokinetics measured in animals. The pharmacokinetic properties of ISIS 301012 provide guidance for clinical development and support infrequent dose administration.

  15. Recombinant human erythropoietin suppresses endothelial cell apoptosis and reduces the ratio of Bax to Bcl-2 proteins in the aortas of apolipoprotein E-deficient mice

    PubMed Central

    Warren, Jeffrey S.; Zhao, Ying; Yung, Raymond; Desai, Anjali

    2013-01-01

    Recent clinical trials have raised concern that therapy with recombinant human erythropoietin (EPO) may increase cardiovascular disease risk, event rate, and mortality. Endothelial cell (EC) apoptosis has been implicated in both atherogenesis as well as in the destabilization and rupture of atheromatous plaques. In the current study we observed that EPO and the EPO-mimetic peptide EMP-1 markedly suppressed lipopolysaccharide-induced apoptosis in EC monolayers. Therapeutic concentrations of EPO upregulated Bcl-2 expression while concurrently diminishing expression of Bax, resulting in a net decrease in the ratio of Bax to Bcl-2 protein concentrations. In vivo studies demonstrated that erythropoietin receptor (EPOR) is abundantly expressed in murine aorta and that EPO treatment for 10 weeks markedly decreased the ratio of Bax to Bcl-2 protein in the aortas of apolipoprotein E-deficient (APO E-KO) mice fed a high-fat diet. To our knowledge these data are the first to reveal a modulation of regulators of the apoptotic pathway in murine aorta by chronic EPO treatment. These observations imply that long-term administration of EPO may have the potential to affect plaque stability. PMID:21242808

  16. Apolipoprotein-E genotypes and myasthenia gravis.

    PubMed

    Suhail, Hamid; Soundararajan, Christhunesa C; Vivekanandhan, Subbiah; Singh, Sumit; Behari, Madhuri

    2010-01-01

    Autoimmune myasthenia gravis (MG) is a disorder of neuromuscular junction. Possible role of multiple genes in the development of the MG has been documented. This case-control study, studied the association of apolipoprotein E (Apo-E) alleles with MG. Anti-AChR antibody was measured using radio receptor immunoassay. Apo-E genotypes were analyzed in 120 MG patients and 120 healthy subjects. Comparison between patients with MG and controls showed no significant association with Apo-E allelic variants. However, a significant association of Apo-E4 allele with AChR-antibody positive patients was observed (P = 0.007). Also, among seropositive patients, a significant association was seen between female gender and Apo-E4 allele (P = 0.023). Our results suggest that the presence of Apo-E4 allele might influence seropositive status in patients with MG and seems an associated susceptible factor in female patients.

  17. Independent effects of apolipoprotein AV and apolipoprotein CIII on plasma triglyceride concentrations

    SciTech Connect

    Baroukh, Nadine N.; Bauge, Eric; Akiyama, Jennifer; Chang, Jessie; Fruchart, Jean-Charles; Rubin, Edward M.; Fruchart, Jamila; Pennacchio, Len A.

    2003-08-15

    Both the apolipoprotein A5 and C3 genes have repeatedly been shown to play an important role in determining plasma triglyceride concentrations in humans and mice. In mice, transgenic and knockout experiments indicate that plasma triglyceride levels are negatively and positively correlated with APOA5 and APOC3 expression, respectively. In humans, common polymorphisms in both genes have also been associated with plasma triglyceride concentrations. The evolutionary relationship among these two apolipoprotein genes and their close proximity on human chromosome 11q23 have largely precluded the determination of their relative contribution to altered Both the apolipoprotein A5 and C3 genes have repeatedly been shown to play an important role in determining plasma triglyceride concentrations in humans and mice. In mice, transgenic and knockout experiments indicate that plasma triglyceride levels are negatively and positively correlated with APOA5 and APOC3 expression, respectively. In humans, common polymorphisms in both genes have also been associated with plasma triglyceride concentrations. The evolutionary relationship among these two apolipoprotein genes and their close proximity on human chromosome 11q23 have largely precluded the determination of their relative contribution to altered triglycerides. To overcome these confounding factors and address their relationship, we generated independent lines of mice that either over-expressed (''double transgenic'') or completely lacked (''double knockout'') both apolipoprotein genes. We report that both ''double transgenic'' and ''double knockout'' mice display intermedia tetriglyceride concentrations compared to over-expression or deletion of either gene alone. Furthermore, we find that human ApoAV plasma protein levels in the ''double transgenic'' mice are approximately 500-fold lower than human ApoCIII levels, supporting ApoAV is a potent triglyceride modulator despite its low concentration. Together, these data indicate

  18. Black knight of AI

    SciTech Connect

    Rose, F.

    1985-03-01

    For two decades now, Hubert Dreyfus, an existentialist philosopher at the University of California at Berkeley, has been in the forefront of the controversy over artificial intelligence. He maintains that computers will never be able to think because scientists will never come up with a suitably rigorous set of rules to describe how we think. To many computer scientists, this is like saying the Earth is flat. But so far, none of them have been able to prove him wrong. Even most AI researchers now admit that before they can make computers any smarter, they'll have to come up with an explanation of how intelligence works in people. This realization has coincided with the emergence of cognitive science, a new discipline linking philosophy, psychology, anthroplogy, linguistics, neuroscience, and computer science in an attempt to develop a theory of the way humans think. The guiding principle of most cognitive science research is the notion that the mind, like the computer, is a system for manipulating symbols - for processing information. The task of cognitive science is to discover how this processing occurs.

  19. T'ai Chi

    MedlinePlus

    ... chi (pronounced: TY CHEE) is great for improving flexibility and strengthening your legs, abs, and arms. What ... general, though, practicing t'ai chi improves strength, flexibility, and respiratory function (breathing). So where can you ...

  20. AI aerospace components

    SciTech Connect

    Heindel, T.A.; Murphy, T.B.; Rasmussen, A.N.; Mcfarland, R.Z.; Montgomery, R.E.; Pohle, G.E.; Heard, A.E.; Atkinson, D.J.; Wedlake, W.E.; Anderson, J.M. Mitre Corp., Houston, TX Unisys Corp., Houston, TX Rockwell International Corp., El Segundo, CA NASA, Kennedy Space Center, Cocoa Beach, FL JPL, Pasadena, CA Lockheed Missiles and Space Co., Inc., Austin, TX McDonnell Douglas Electronic Systems Co., McLean, VA )

    1991-10-01

    An evaluation is made of the application of novel, AI-capabilities-related technologies to aerospace systems. Attention is given to expert-system shells for Space Shuttle Orbiter mission control, manpower and processing cost reductions at the NASA Kennedy Space Center's 'firing rooms' for liftoff monitoring, the automation of planetary exploration systems such as semiautonomous mobile robots, and AI for battlefield staff-related functions.

  1. AI aerospace components

    NASA Technical Reports Server (NTRS)

    Heindel, Troy A.; Murphy, Terri B.; Rasmussen, Arthur N.; Mcfarland, Robert Z.; Montgomery, Ronnie E.; Pohle, George E.; Heard, Astrid E.; Atkinson, David J.; Wedlake, William E.; Anderson, John M.

    1991-01-01

    An evaluation is made of the application of novel, AI-capabilities-related technologies to aerospace systems. Attention is given to expert-system shells for Space Shuttle Orbiter mission control, manpower and processing cost reductions at the NASA Kennedy Space Center's 'firing rooms' for liftoff monitoring, the automation of planetary exploration systems such as semiautonomous mobile robots, and AI for battlefield staff-related functions.

  2. AI aerospace components

    NASA Technical Reports Server (NTRS)

    Heindel, Troy A.; Murphy, Terri B.; Rasmussen, Arthur N.; Mcfarland, Robert Z.; Montgomery, Ronnie E.; Pohle, George E.; Heard, Astrid E.; Atkinson, David J.; Wedlake, William E.; Anderson, John M.

    1991-01-01

    An evaluation is made of the application of novel, AI-capabilities-related technologies to aerospace systems. Attention is given to expert-system shells for Space Shuttle Orbiter mission control, manpower and processing cost reductions at the NASA Kennedy Space Center's 'firing rooms' for liftoff monitoring, the automation of planetary exploration systems such as semiautonomous mobile robots, and AI for battlefield staff-related functions.

  3. Artificial intelligence. Fears of an AI pioneer.

    PubMed

    Russell, Stuart; Bohannon, John

    2015-07-17

    From the enraged robots in the 1920 play R.U.R. to the homicidal computer H.A.L. in 2001: A Space Odyssey, science fiction writers have embraced the dark side of artificial intelligence (AI) ever since the concept entered our collective imagination. Sluggish progress in AI research, especially during the “AI winter” of the 1970s and 1980s, made such worries seem far-fetched. But recent breakthroughs in machine learning and vast improvements in computational power have brought a flood of research fun