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Sample records for cholesterol transport proteins

  1. Overexpression and deletion of phospholipid transfer protein reduce HDL mass and cholesterol efflux capacity but not macrophage reverse cholesterol transport[S

    Science.gov (United States)

    Kuwano, Takashi; Bi, Xin; Cipollari, Eleonora; Yasuda, Tomoyuki; Lagor, William R.; Szapary, Hannah J.; Tohyama, Junichiro; Millar, John S.; Billheimer, Jeffrey T.; Lyssenko, Nicholas N.; Rader, Daniel J.

    2017-01-01

    Phospholipid transfer protein (PLTP) may affect macrophage reverse cholesterol transport (mRCT) through its role in the metabolism of HDL. Ex vivo cholesterol efflux capacity and in vivo mRCT were assessed in PLTP deletion and PLTP overexpression mice. PLTP deletion mice had reduced HDL mass and cholesterol efflux capacity, but unchanged in vivo mRCT. To directly compare the effects of PLTP overexpression and deletion on mRCT, human PLTP was overexpressed in the liver of wild-type animals using an adeno-associated viral (AAV) vector, and control and PLTP deletion animals were injected with AAV-null. PLTP overexpression and deletion reduced plasma HDL mass and cholesterol efflux capacity. Both substantially decreased ABCA1-independent cholesterol efflux, whereas ABCA1-dependent cholesterol efflux remained the same or increased, even though preβ HDL levels were lower. Neither PLTP overexpression nor deletion affected excretion of macrophage-derived radiocholesterol in the in vivo mRCT assay. The ex vivo and in vivo assays were modified to gauge the rate of cholesterol efflux from macrophages to plasma. PLTP activity did not affect this metric. Thus, deviations in PLTP activity from the wild-type level reduce HDL mass and ex vivo cholesterol efflux capacity, but not the rate of macrophage cholesterol efflux to plasma or in vivo mRCT. PMID:28137768

  2. How cholesterol interacts with proteins and lipids during its intracellular transport

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Solanko, Katarzyna

    2015-01-01

    Sterols, as cholesterol in mammalian cells and ergosterol in fungi, are indispensable molecules for proper functioning and nanoscale organization of the plasma membrane. Synthesis, uptake and efflux of cholesterol are regulated by a variety of protein-lipid and protein-protein interactions. Simil...... specific protein-lipid and protein-protein interactions help overcoming the extremely low water solubility of cholesterol, thereby controlling intracellular cholesterol movement. This article is part of a Special Issue entitled: Lipid-protein interactions.......Sterols, as cholesterol in mammalian cells and ergosterol in fungi, are indispensable molecules for proper functioning and nanoscale organization of the plasma membrane. Synthesis, uptake and efflux of cholesterol are regulated by a variety of protein-lipid and protein-protein interactions....... Similarly, membrane lipids and their physico-chemical properties directly affect cholesterol partitioning and thereby contribute to the highly heterogeneous intracellular cholesterol distribution. Movement of cholesterol in cells is mediated by vesicle trafficking along the endocytic and secretory pathways...

  3. The endoplasmic reticulum coat protein II transport machinery coordinates cellular lipid secretion and cholesterol biosynthesis

    NARCIS (Netherlands)

    Fryer, Lee G. D.; Jones, Bethan; Duncan, Emma J.; Hutchison, Claire E.; Ozkan, Tozen; Williams, Paul A.; Alder, Olivia; Nieuwdorp, Max; Townley, Anna K.; Mensenkamp, Arjen R.; Stephens, David J.; Dallinga-Thie, Geesje M.; Shoulders, Carol C.

    2014-01-01

    Triglycerides and cholesterol are essential for life in most organisms. Triglycerides serve as the principal energy storage depot and, where vascular systems exist, as a means of energy transport. Cholesterol is essential for the functional integrity of all cellular membrane systems. The endoplasmic

  4. Pitavastatin Differentially Modulates MicroRNA-Associated Cholesterol Transport Proteins in Macrophages.

    Directory of Open Access Journals (Sweden)

    Haijun Zhang

    Full Text Available There is emerging evidence identifying microRNAs (miRNAs as mediators of statin-induced cholesterol efflux, notably through the ATP-binding cassette transporter A1 (ABCA1 in macrophages. The objective of this study was to assess the impact of an HMG-CoA reductase inhibitor, pitavastatin, on macrophage miRNAs in the presence and absence of oxidized-LDL, a hallmark of a pro-atherogenic milieu. Treatment of human THP-1 cells with pitavastatin prevented the oxLDL-mediated suppression of miR-33a, -33b and -758 mRNA in these cells, an effect which was not uniquely attributable to induction of SREBP2. Induction of ABCA1 mRNA and protein by oxLDL was inhibited (30% by pitavastatin, while oxLDL or pitavastatin alone significantly induced and repressed ABCA1 expression, respectively. These findings are consistent with previous reports in macrophages. miRNA profiling was also performed using a miRNA array. We identified specific miRNAs which were up-regulated (122 and down-regulated (107 in THP-1 cells treated with oxLDL plus pitavastatin versus oxLDL alone, indicating distinct regulatory networks in these cells. Moreover, several of the differentially expressed miRNAs identified are functionally associated with cholesterol trafficking (six miRNAs in cells treated with oxLDL versus oxLDL plus pitavastatin. Our findings indicate that pitavastatin can differentially modulate miRNA in the presence of oxLDL; and, our results provide evidence that the net effect on cholesterol homeostasis is mediated by a network of miRNAs.

  5. Role of Heat Shock Factor 1 in Conserving Cholesterol Transportation in Leydig Cell Steroidogenesis via Steroidogenic Acute Regulatory Protein.

    Science.gov (United States)

    Oka, Shintaro; Shiraishi, Koji; Fujimoto, Mitsuaki; Katiyar, Arpit; Takii, Ryosuke; Nakai, Akira; Matsuyama, Hideyasu

    2017-08-01

    Testicular testosterone synthesis begins with cholesterol transport into mitochondria via steroidogenic acute regulatory (StAR) protein in Leydig cells. Acute heat stress is known to obstruct testicular steroidogenesis by transcriptional repression of StAR. In contrast, chronic heat stress such as cryptorchidism or varicocele generally does not affect testicular steroidogenesis, suggesting that Leydig cells adapt to heat stress and retain their steroid synthesis ability. However, the mechanisms of the stress response in steroid-producing cells are unclear. We examined the relationship between the heat stress response and heat shock factor 1 (HSF1), which protects cells from proteotoxic stress by inducing heat shock protein as a molecular chaperone. The influences of HSF1 deficiency on cholesterol transport by StAR and the expression of steroidogenic enzymes under chronic heat stress were studied in testes of HSF1-knockout (HSF1KO) mice with experimental cryptorchidism. StAR protein in wild-type-cryptorchid mice was transiently decreased after induction of cryptorchidism and then gradually returned to basal levels. In contrast, StAR protein in HSF1KO mice continued to decrease and failed to recover, resulting in impaired serum testosterone. StAR messenger RNA was not decreased with cryptorchidism, indicating that posttranslational modification of StAR, not its transcription, was obstructed in cryptorchidism. Other steroidogenic enzymes, including CYP11A1, 3β-HSD, and CYP17A1, were not decreased. Lipid droplets were increased in the cytosol of HSF1KO-cryptorchid mice, suggesting dysfunctional cholesterol transportation. These findings provide insight into the role of HSF1 in Leydig cell steroidogenesis, suggesting that it maintains cholesterol transport by recovering StAR under chronic heat stress. Copyright © 2017 Endocrine Society.

  6. Maternal-fetal cholesterol transport in the second half of mouse pregnancy does not involve LDL receptor-related protein 2.

    Science.gov (United States)

    Zwier, M V; Baardman, M E; van Dijk, T H; Jurdzinski, A; Wisse, L J; Bloks, V W; Berger, R M F; DeRuiter, M C; Groen, A K; Plösch, T

    2017-08-01

    LDL receptor-related protein type 2 (LRP2) is highly expressed on both yolk sac and placenta. Mutations in the corresponding gene are associated with severe birth defects in humans, known as Donnai-Barrow syndrome. We here characterized the contribution of LRP2 and maternal plasma cholesterol availability to maternal-fetal cholesterol transport and fetal cholesterol levels in utero in mice. Lrp2 +/- mice were mated heterozygously to yield fetuses of all three genotypes. Half of the dams received a 0.5% probucol-enriched diet during gestation to decrease maternal HDL cholesterol. At E13.5, the dams received an injection of D7-labelled cholesterol and were provided with 1- 13 C acetate-supplemented drinking water. At E16.5, fetal tissues were collected and maternal cholesterol transport and fetal synthesis quantified by isotope enrichments in fetal tissues by GC-MS. The Lrp2 genotype did not influence maternal-fetal cholesterol transport and fetal cholesterol. However, lowering of maternal plasma cholesterol levels by probucol significantly reduced maternal-fetal cholesterol transport. In the fetal liver, this was associated with increased cholesterol synthesis rates. No indications were found for an interaction between the Lrp2 genotype and maternal probucol treatment. Maternal-fetal cholesterol transport and endogenous fetal cholesterol synthesis depend on maternal cholesterol concentrations but do not involve LRP2 in the second half of murine pregnancy. Our results suggest that the mouse fetus can compensate for decreased maternal cholesterol levels. It remains a relevant question how the delicate system of cholesterol transport and synthesis is regulated in the human fetus and placenta. © 2016 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

  7. Characterization of placental cholesterol transport

    DEFF Research Database (Denmark)

    Lindegaard, Marie L; Wassif, Christopher A; Vaisman, Boris

    2008-01-01

    Patients with Smith-Lemli-Opitz syndrome (SLOS) are born with multiple congenital abnormalities. Postnatal cholesterol supplementation is provided; however, it cannot correct developmental malformations due to in utero cholesterol deficit. Increased transport of cholesterol from maternal to fetal...... circulation might attenuate congenital malformations. The cholesterol transporters Abca1, Abcg1, and Sr-b1 are present in placenta; however, their potential role in placental transport remains undetermined. In mice, expression analyses showed that Abca1 and Abcg1 transcripts increased 2-3-fold between...... embryonic days 13.5 and 18.5 in placental tissue; whereas, Sr-b1 expression decreased. To examine the functional role of Abca1, Abcg1 and Sr-b1 we measured the maternal-fetal transfer of (14)C-cholesterol in corresponding mutant embryos. Disruption of either Abca1 or Sr-b1 decreased cholesterol transfer...

  8. Relevance of hereditary defects in lipid transport proteins for the pathogenesis of cholesterol gallstone disease

    NARCIS (Netherlands)

    vanBerge-Henegouwen, G. P.; Venneman, N. G.; Portincasa, P.; Kosters, A.; van Erpecum, K. J.; Groen, A. K.

    2004-01-01

    In the formation of cholesterol gallstones, cholesterol hypersecretion into bile causing cholesterol supersaturation and crystallization appears to be the primary factor, with disturbed gallbladder and intestinal motility as secondary factors. Although intestinal uptake mechanisms have not yet been

  9. Synthetic High-Density Lipoprotein-Like Nanocarrier Improved Cellular Transport of Lysosomal Cholesterol in Human Sterol Carrier Protein-Deficient Fibroblasts.

    Science.gov (United States)

    Nam, Da-Eun; Kim, Ok-Kyung; Park, Yoo Kyoung; Lee, Jeongmin

    2016-01-01

    Sterol carrier protein-2 (SCP-2), which is not found in tissues of people with Zellweger syndrome, facilitates the movement of cholesterol within cells, resulting in abnormal accumulation of cholesterol in SCP-2-deficient cells. This study investigated whether synthetic high-density lipoprotein-like nanocarrier (sHDL-NC) improves the cellular transport of lysosomal cholesterol to plasma membrane in SCP-2-deficient fibroblasts. Human SCP-2-deficient fibroblasts were incubated with [(3)H-cholesterol]LDL as a source of cholesterol and sHDL-NC. The cells were fractionated by centrifugation permit tracking of [(3)H]-cholesterol from lysosome into plasma membrane. Furthermore, cellular content of cholesteryl ester as a storage form and mRNA expression of low-density lipoprotein (LDL) receptor were measured to support the cholesterol transport to plasma membrane. Incubation with sHDL-NC for 8 h significantly increased uptake of [(3)H]-cholesterol to lysosome by 53% and further enhanced the transport of [(3)H]-cholesterol to plasma membrane by 32%. Treatment with sHDL-NC significantly reduced cellular content of cholesteryl ester and increased mRNA expression of LDL receptor (LDL-R). In conclusion, sHDL-NC enables increased transport of lysosomal cholesterol to plasma membrane. In addition, these data were indirectly supported by decreased cellular content of cholesteryl ester and increased gene expression of LDL-R. Therefore, sHDL-NC may be a useful vehicle for transporting cholesterol, which may help to prevent accumulation of cholesterol in SCP-2-deficient fibroblasts.

  10. Lupin protein isolate versus casein modifies cholesterol excretion and mRNA expression of intestinal sterol transporters in a pig model.

    Science.gov (United States)

    Radtke, Juliane; Geissler, Stefanie; Schutkowski, Alexandra; Brandsch, Corinna; Kluge, Holger; Duranti, Marcello M; Keller, Sylvia; Jahreis, Gerhard; Hirche, Frank; Stangl, Gabriele I

    2014-02-03

    Lupin proteins exert hypocholesterolemic effects in man and animals, although the underlying mechanism remains uncertain. Herein we investigated whether lupin proteins compared to casein modulate sterol excretion and mRNA expression of intestinal sterol transporters by use of pigs as an animal model with similar lipid metabolism as humans, and cellular cholesterol-uptake by Caco-2 cells. Two groups of pigs were fed cholesterol-containing diets with either 230 g/kg of lupin protein isolate from L. angustifolius or 230 g/kg casein, for 4 weeks. Faeces were collected quantitatively over a 5 d period for analysis of neutral sterols and bile acids by gas chromatographically methods. The mRNA abundances of intestinal lipid transporters were analysed by real-time RT-PCR. Cholesterol-uptake studies were performed with Caco-2 cells that were incubated with lupin conglutin γ, phytate, ezetimibe or albumin in the presence of labelled [4-14C]-cholesterol. Pigs fed the lupin protein isolate revealed lower cholesterol concentrations in total plasma, LDL and HDL than pigs fed casein (P isolate compared to pigs that received casein (+57.1%; P isolate than in those who received casein (P isolate is attributable to an increased faecal output of cholesterol and a reduced intestinal uptake of cholesterol. The findings indicate phytate as a possible biofunctional ingredient of lupin protein isolate.

  11. Intestinal cholesterol transport: Measuring cholesterol absorption and its reverse

    NARCIS (Netherlands)

    Jakulj, L.

    2013-01-01

    Intestinal cholesterol transport might serve as an attractive future target for cardiovascular disease reduction, provided that underlying molecular mechanisms are more extensively elucidated, combined with improved techniques to measure changes in cholesterol fluxes and their possible

  12. ABC-transporters and lipid transfer proteins : important players in macrophage cholesterol homeostasis and atherosclerosis

    NARCIS (Netherlands)

    Ye, Dan

    2008-01-01

    Local modulation of macrophage cholesterol metabolism in the arterial wall and systemic regulation of lipoprotein metabolism (LDL-lowering and/or HDL-raising) are both attractive targets for future drug design for the prevention of atherosclerosis. As described in this thesis, bone marrow

  13. Maternal-fetal cholesterol transport in the second half ofmouse pregnancy does not involve LDL receptor-related protein 2

    NARCIS (Netherlands)

    Zwier, Mathijs V; Baardman, Maria E; van Dijk, Theo H; Jurdzinski, Angelika; Wisse, Lambertus J; Bloks, Vincent W; Berger, Rolf M F; DeRuiter, Marco C; Groen, Albert K; Plösch, Torsten

    AimLDL receptor-related protein type 2 (LRP2) is highly expressed on both yolk sac and placenta. Mutations in the corresponding gene are associated with severe birth defects in humans, known as Donnai-Barrow syndrome. We here characterized the contribution of LRP2 and maternal plasma cholesterol

  14. Cellular Cholesterol Regulates Ubiquitination and Degradation of the Cholesterol Export Proteins ABCA1 and ABCG1*

    Science.gov (United States)

    Hsieh, Victar; Kim, Mi-Jurng; Gelissen, Ingrid C.; Brown, Andrew J.; Sandoval, Cecilia; Hallab, Jeannette C.; Kockx, Maaike; Traini, Mathew; Jessup, Wendy; Kritharides, Leonard

    2014-01-01

    The objective of this study was to examine the influence of cholesterol in post-translational control of ABCA1 and ABCG1 protein expression. Using CHO cell lines stably expressing human ABCA1 or ABCG1, we observed that the abundance of these proteins is increased by cell cholesterol loading. The response to increased cholesterol is rapid, is independent of transcription, and appears to be specific for these membrane proteins. The effect is mediated through cholesterol-dependent inhibition of transporter protein degradation. Cell cholesterol loading similarly regulates degradation of endogenously expressed ABCA1 and ABCG1 in human THP-1 macrophages. Turnover of ABCA1 and ABCG1 is strongly inhibited by proteasomal inhibitors and is unresponsive to inhibitors of lysosomal proteolysis. Furthermore, cell cholesterol loading inhibits ubiquitination of ABCA1 and ABCG1. Our findings provide evidence for a rapid, cholesterol-dependent, post-translational control of ABCA1 and ABCG1 protein levels, mediated through a specific and sterol-sensitive mechanism for suppression of transporter protein ubiquitination, which in turn decreases proteasomal degradation. This provides a mechanism for acute fine-tuning of cholesterol transporter activity in response to fluctuations in cell cholesterol levels, in addition to the longer term cholesterol-dependent transcriptional regulation of these genes. PMID:24500716

  15. Scavenger receptor class B member 1 protein: hepatic regulation and its effects on lipids, reverse cholesterol transport, and atherosclerosis

    Directory of Open Access Journals (Sweden)

    Kent AP

    2011-04-01

    Full Text Available Anthony P Kent, Ioannis M StylianouDepartment of Medicine and Institute for Translational Medicine and Therapeutics, University of Pennsylvania School of Medicine, Philadelphia, PA, USAAbstract: Scavenger receptor class B member 1 (SR-BI, also known as SCARB1 is the primary receptor for the selective uptake of cholesterol from high-density lipoprotein (HDL. SR-BI is present in several key tissues; however, its presence and function in the liver is deemed the most relevant for protection against atherosclerosis. Cholesterol is transferred from HDL via SR-BI to the liver, which ultimately results in the excretion of cholesterol via bile and feces in what is known as the reverse cholesterol transport pathway. Much of our knowledge of SR-BI hepatic function and regulation is derived from mouse models and in vitro characterization. Multiple independent regulatory mechanisms of SR-BI have been discovered that operate at the transcriptional and post-transcriptional levels. In this review we summarize the critical discoveries relating to hepatic SR-BI cholesterol metabolism, atherosclerosis, and regulation of SR-BI, as well as alternative functions that may indirectly affect atherosclerosis.Keywords: SR-BI, SCARB1, lipids, atherosclerosis, CAD, mouse models

  16. Alterations in high-density lipoprotein metabolism and reverse cholesterol transport in insulin resistance and type 2 diabetes mellitus : role of lipolytic enzymes, lecithin : cholesterol acyltransferase and lipid transfer proteins

    NARCIS (Netherlands)

    Borggreve, SE; de Vries, R; Dullaart, RPF

    2003-01-01

    Insulin resistance and type 2 diabetes mellitus are generally accompanied by low HDL cholesterol and high plasma triglycerides, which are major cardiovascular risk factors. This review describes abnormalities in HDL metabolism and reverse cholesterol transport, i.e. the transport of cholesterol from

  17. Intracellular transport of cholesterol in mammalian cells

    International Nuclear Information System (INIS)

    Brasaemle, D.L.

    1989-01-01

    The erythrocyte was selected as a simple cell for the study of transbilayer movement of cholesterol. Cholesterol oxidase was used to measure the distribution of [ 3 H]cholesterol across the erythrocyte membrane. Cholesterol oxidase was also used to estimate the rate of transport of low density lipoprotein (LDL) cholesterol to the plasma membrane of cultured Chinese hamster ovary (CHO) fibroblasts; the half-time of this process was 42 minutes. The rate of transport of LDL cholesterol to the plasma membrane was confirmed by a second procedure using amphotericin B. Amphotericin B was also used to estimate the rate of transport of endogenously synthesized cholesterol to the plasma membrane of CHO cells. New methodology was developed including improvements of the previously published cholesterol oxidase assay for plasma membrane cholesterol. A new method for detecting transport of cholesterol to the plasma membrane in cultured cells was developed using amphotericin B. Preliminary studies investigated the use of fluorescent polyenes, pimaricin and etruscomycin, as probes for plasma membrane cholesterol in transport studies. Finally, a modification of a previously published cell staining protocol yielded a simple, quantitative assay for cell growth

  18. Human paraoxonase 1 overexpression in mice stimulates HDL cholesterol efflux and reverse cholesterol transport

    Science.gov (United States)

    Ikhlef, Souade; Berrougui, Hicham; Kamtchueng Simo, Olivier; Zerif, Echarki

    2017-01-01

    This study was aimed to investigate the effect of human PON1 overexpression in mice on cholesterol efflux and reverse cholesterol transport. PON1 overexpression in PON1-Tg mice induced a significant 3-fold (pparaoxonase activity and a significant ~30% (p<0.0001) increase in the capacity of HDL to mediate cholesterol efflux from J774 macrophages compared to wild-type mice. It also caused a significant 4-fold increase (p<0.0001) in the capacity of macrophages to transfer cholesterol to apoA-1, a significant 2-fold (p<0.0003) increase in ABCA1 mRNA and protein expression, and a significant increase in the expression of PPARγ (p<0.0003 and p<0.04, respectively) and LXRα (p<0.0001 and p<0.01, respectively) mRNA and protein compared to macrophages from wild-type mice. Moreover, transfection of J774 macrophages with human PON1 also increased ABCA1, PPARγ and LXRα protein expression and stimulates macrophages cholesterol efflux to apo A1. In vivo measurements showed that the overexpression of PON1 significantly increases the fecal elimination of macrophage-derived cholesterol in PON1-Tg mice. Overall, our results suggested that the overexpression of PON1 in mice may contribute to the regulation of the cholesterol homeostasis by improving the capacity of HDL to mediate cholesterol efflux and by stimulating reverse cholesterol transport. PMID:28278274

  19. Upregulating reverse cholesterol transport with cholesteryl ester transfer protein inhibition requires combination with the LDL-lowering drug berberine in dyslipidemic hamsters.

    Science.gov (United States)

    Briand, François; Thieblemont, Quentin; Muzotte, Elodie; Sulpice, Thierry

    2013-01-01

    This study aimed to investigate whether cholesteryl ester transfer protein inhibition promotes in vivo reverse cholesterol transport in dyslipidemic hamsters. In vivo reverse cholesterol transport was measured after an intravenous injection of (3)H-cholesteryl-oleate-labeled/oxidized low density lipoprotein particles ((3)H-oxLDL), which are rapidly cleared from plasma by liver-resident macrophages for further (3)H-tracer egress in plasma, high density lipoprotein (HDL), liver, and feces. A first set of hamsters made dyslipidemic with a high-fat and high-fructose diet was treated with vehicle or torcetrapib 30 mg/kg (TOR) over 2 weeks. Compared with vehicle, TOR increased apolipoprotein E-rich HDL levels and significantly increased (3)H-tracer appearance in HDL by 30% over 72 hours after (3)H-oxLDL injection. However, TOR did not change (3)H-tracer recovery in liver and feces, suggesting that uptake and excretion of cholesterol deriving from apolipoprotein E-rich HDL is not stimulated. As apoE is a potent ligand for the LDL receptor, we next evaluated the effects of TOR in combination with the LDL-lowering drug berberine, which upregulates LDL receptor expression in dyslipidemic hamsters. Compared with TOR alone, treatment with TOR+berberine 150 mg/kg resulted in lower apolipoprotein E-rich HDL levels. After (3)H-oxLDL injection, TOR+berberine significantly increased (3)H-tracer appearance in fecal cholesterol by 109%. Our data suggest that cholesteryl ester transfer protein inhibition alone does not stimulate reverse cholesterol transport in dyslipidemic hamsters and that additional effects mediated by the LDL-lowering drug berberine are required to upregulate this process.

  20. Comparative effect of fish oil feeding and other dietary fatty acids on plasma lipoproteins, biliary lipids, and hepatic expression of proteins involved in reverse cholesterol transport in the rat.

    Science.gov (United States)

    Morgado, Nora; Rigotti, Attilio; Valenzuela, Alfonso

    2005-01-01

    While elevated plasma high-density lipoprotein (HDL) levels has been associated to a reduction in cardiovascular risk, dietary fish oils rich in omega-3 polyunsaturated fatty acids (PUFAs) may protect against this disease. The protective effect of HDL is associated to its participation in the reverse cholesterol transport pathway. On the other hand, omega-3 PUFAs decrease plasma HDL levels compared to other fatty acids, which may suggest an effect on reverse cholesterol transport. In this work, the effect of dietary fish oil on the fatty acid composition of hepatic membranes, plasma lipoprotein cholesterol profile, biliary lipids, and the expression of proteins involved in reverse cholesterol transport, was compared to other dietary oils having a different degree of fatty acid unsaturation. Male rats were fed a semi synthetic diet containing fish oil (omega-3), sunflower oil (omega-6), olive oil (omega-9) or coconut oil (saturated). Hepatic membrane fatty acid composition, plasma cholesterol levels, lipoprotein cholesterol profile, biliary lipids, hepatic mRNA levels for lecithin cholesterol acyltransferase, hepatic lipase, apo E, and apo A-I, and hepatic protein levels of the scavenger receptor class B type I, caveolin-1, and the ATP binding cassette transporter A1 were analyzed. Plasma apo A-I and apo E protein levels were also evaluated. Compared to the other diets, omega-3 PUFAs significantly changed omega-3/omega-6 fatty acid ratio of hepatic membranes, caused a reduction of plasma total and HDL cholesterol, and selectively increased biliary cholesterol secretion. No modification in the expression levels of lecithin cholesterol acyltransferase, hepatic lipase, apo A-I and apo E mRNA was observed. Hepatic scavenger receptor class B type I, caveolin-1, and the ATP binding cassette transporter A1 protein levels were also not affected. Plasma apo A-I, but not apo E, was reduced. These results show that dietary omega-3 PUFAs reduce plasma HDL cholesterol and

  1. Alcohol consumption stimulates early steps in reverse cholesterol transport

    OpenAIRE

    Gaag, M.S. van der; Tol, A. van; Vermunt, S.H.F.; Scheek, L.M.; Schaafsma, G.; Hendriks, H.F.J.

    2001-01-01

    Alcohol consumption is associated with increased HDL cholesterol levels, which may indicate stimulated reverse cholesterol transport. The mechanism is, however, not known. The aim of this study was to evaluate the effects of alcohol consumption on the first two steps of the reverse cholesterol pathway: cellular cholesterol efflux and plasma cholesterol esterification. Eleven healthy middle-aged men consumed four glasses (40 g of alcohol) of red wine, beer, spirits (Dutch gin), or carbonated m...

  2. The Role of Macrophage Lipophagy in Reverse Cholesterol Transport

    Directory of Open Access Journals (Sweden)

    Se-Jin Jeong

    2017-03-01

    Full Text Available Macrophage cholesterol efflux is a central step in reverse cholesterol transport, which helps to maintain cholesterol homeostasis and to reduce atherosclerosis. Lipophagy has recently been identified as a new step in cholesterol ester hydrolysis that regulates cholesterol efflux, since it mobilizes cholesterol from lipid droplets of macrophages via autophagy and lysosomes. In this review, we briefly discuss recent advances regarding the mechanisms of the cholesterol efflux pathway in macrophage foam cells, and present lipophagy as a therapeutic target in the treatment of atherosclerosis.

  3. Cholesterol Transport Revisited: A New Turbo Mechanism to Drive Cholesterol Excretion

    NARCIS (Netherlands)

    de Boer, Jan Freark; Kuipers, Folkert; Groen, Albert K.

    2018-01-01

    A fine-tuned balance between cholesterol uptake and excretion by the body is pivotal to maintain health and to remain free from the deleterious consequences of cholesterol accumulation such as cardiovascular disease. The pathways involved in intracellular and extracellular cholesterol transport are

  4. Cholesterol Transport Revisited : A New Turbo Mechanism to Drive Cholesterol Excretion

    NARCIS (Netherlands)

    de Boer, Jan Freark; Kuipers, Folkert; Groen, Albert K.

    A fine-tuned balance between cholesterol uptake and excretion by the body is pivotal to maintain health and to remain free from the deleterious consequences of cholesterol accumulation such as cardiovascular disease. The pathways involved in intracellular and extracellular cholesterol transport are

  5. Continuous transport of a small fraction of plasma membrane cholesterol to endoplasmic reticulum regulates total cellular cholesterol.

    Science.gov (United States)

    Infante, Rodney Elwood; Radhakrishnan, Arun

    2017-04-17

    Cells employ regulated transport mechanisms to ensure that their plasma membranes (PMs) are optimally supplied with cholesterol derived from uptake of low-density lipoproteins (LDL) and synthesis. To date, all inhibitors of cholesterol transport block steps in lysosomes, limiting our understanding of post-lysosomal transport steps. Here, we establish the cholesterol-binding domain 4 of anthrolysin O (ALOD4) as a reversible inhibitor of cholesterol transport from PM to endoplasmic reticulum (ER). Using ALOD4, we: (1) deplete ER cholesterol without altering PM or overall cellular cholesterol levels; (2) demonstrate that LDL-derived cholesterol travels from lysosomes first to PM to meet cholesterol needs, and subsequently from PM to regulatory domains of ER to suppress activation of SREBPs, halting cholesterol uptake and synthesis; and (3) determine that continuous PM-to-ER cholesterol transport allows ER to constantly monitor PM cholesterol levels, and respond rapidly to small declines in cellular cholesterol by activating SREBPs, increasing cholesterol uptake and synthesis.

  6. Protein complexes and cholesterol in the control of late endosomal dynamicsCholesterol and multi-protein complexes in the control of late endosomal dynamics

    NARCIS (Netherlands)

    Kant, Rik Henricus Nicolaas van der

    2013-01-01

    Late endosomal transport is disrupted in several diseases such as Niemann-Pick type C, ARC syndrome and Alzheimer’s disease. This thesis describes the regulation of late endosomal dynamics by cholesterol and multi-protein complexes. We find that cholesterol acts as a cellular tomtom that steers the

  7. Regulation of biliary cholesterol secretion and reverse cholesterol transport

    NARCIS (Netherlands)

    Dikkers, Arne

    2016-01-01

    According to the World Health Organization the number one cause of death throughout the world is cardiovascular disease. Therefore, there is an urgent need for new therapeutic strategies to prevent and treat cardiovascular disease. One possible way is to target the HDL-driven reverse cholesterol

  8. Alcohol consumption stimulates early stemps in reverse cholesterol transport

    NARCIS (Netherlands)

    Gaag, van der M.S.; Tol, van A.; Vermunt, S.H.F.; Scheek, L.M.; Schaafsma, G.; Hendriks, H.F.J.

    2001-01-01

    Alcohol consumption is associated with increased HDL cholesterol levels, which may indicate stimulated reverse cholesterol transport. The mechanism is, however, not known. The aim of this study was to evaluate the effects of alcohol consumption on the first two steps of the reverse cholesterol

  9. Alcohol consumption stimulates early steps in reverse cholesterol transport

    NARCIS (Netherlands)

    Gaag, M.S. van der; Tol, A. van; Vermunt, S.H.F.; Scheek, L.M.; Schaafsma, G.; Hendriks, H.F.J.

    2001-01-01

    Alcohol consumption is associated with increased HDL cholesterol levels, which may indicate stimulated reverse cholesterol transport. The mechanism is, however, not known. The aim of this study was to evaluate the effects of alcohol consumption on the first two steps of the reverse cholesterol

  10. Impaired reverse cholesterol transport and hepatic steatosis ...

    African Journals Online (AJOL)

    ... relative liver weight, serum lipid profile, expressions of hepatic marker gene of lipid metabolism and liver morphology were observed in three hyperlipidemic models. Results: Elevated total cholesterol (TC), triglyceride, low density lipoprotein-cholesterol (LDL-C) and high density lipoprotein-cholesterol (HDL-C) levels and ...

  11. Effect of Moderate Alcohol Consumption on Parameters of Reverse Cholesterol Transport in Postmenopausal Women

    NARCIS (Netherlands)

    Sierksma, A.; Vermunt, S.H.F.; Lankhuizen, I.M.; Gaag, M.S. van der; Scheek, L.M.; Grobbee, D.E.; Tol, A. van; Hendriks, H.F.J.

    2004-01-01

    Background: Alcohol consumption is associated with increased high-density lipoprotein (HDL) cholesterol levels. One of the main antiatherogenic functions of HDL is reverse cholesterol transport. Three early steps of reverse cholesterol transport are (1) cellular cholesterol efflux, (2) plasma

  12. The GARP Complex Is Involved in Intracellular Cholesterol Transport via Targeting NPC2 to Lysosomes.

    Science.gov (United States)

    Wei, Jian; Zhang, Ying-Yu; Luo, Jie; Wang, Ju-Qiong; Zhou, Yu-Xia; Miao, Hong-Hua; Shi, Xiong-Jie; Qu, Yu-Xiu; Xu, Jie; Li, Bo-Liang; Song, Bao-Liang

    2017-06-27

    Proper intracellular cholesterol trafficking is critical for cellular function. Two lysosome-resident proteins, NPC1 and NPC2, mediate the egress of low-density lipoprotein-derived cholesterol from lysosomes. However, other proteins involved in this process remain largely unknown. Through amphotericin B-based selection, we isolated two cholesterol transport-defective cell lines. Subsequent whole-transcriptome-sequencing analysis revealed two cell lines bearing the same mutation in the vacuolar protein sorting 53 (Vps53) gene. Depletion of VPS53 or other subunits of the Golgi-associated retrograde protein (GARP) complex impaired NPC2 sorting to lysosomes and caused cholesterol accumulation. GARP deficiency blocked the retrieval of the cation-independent mannose 6-phosphate receptor (CI-MPR) to the trans-Golgi network. Further, Vps54 mutant mice displayed reduced cellular NPC2 protein levels and increased cholesterol accumulation, underscoring the physiological role of the GARP complex in cholesterol transport. We conclude that the GARP complex contributes to intracellular cholesterol transport by targeting NPC2 to lysosomes in a CI-MPR-dependent manner. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  13. 27-Hydroxycholesterol regulates cholesterol synthesis and transport in C6 glioma cells.

    Science.gov (United States)

    An, Yu; Zhang, Dan-Di; Yu, Huan-Ling; Ma, Wei-Wei; Lu, Yan-Hui; Liu, Quan-Ri; Xiao, Rong

    2017-03-01

    The oxysterol 27-Hydroxycholesterol (27-OHC) is a major cholesterol metabolite that can cross the blood brain barrier (BBB) from peripheral circulation to the brain. Currently, the role of 27-OHC on cholesterol homeostasis in astrocytes and the underlying mechanisms are not defined. Since all brain cholesterol is essentially synthesized in brain itself and astrocytes as net producers of cholesterol are essential for normal brain function, here we investigated the effects of 27-OHC on cholesterol synthesis and transport in C6 glioma cells. C6 cells were treated with 5, 10 and 20μM 27-OHC for 24h and the cell viability and apoptosis, the cholesterol levels and metabolism-related mediators, genes and proteins were subsequently assessed using cell-counting kit (CCK)-8, Amplex red, ELISA, real-time PCR and Western blot, respectively. We found that 27-OHC decreased cholesterol levels by down-regulating the expression of sterol-regulated element binding protein-1 (SREBP-1a), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CR) and low density lipoprotein receptor (LDLR) and promoted cholesterol transport by up-regulating the expression of peroxisome proliferator-activated receptors-γ (PPAR-γ), liver X receptor-α (LXR-α), ATP-binding cassette transporter protein family member A1 (ABCA1) and apolipoprotein E (ApoE)genes. Our results suggested that 27-OHC may represent a sensitive modulator of cholesterol metabolism disorder by suppressing cholesterol synthesis and stimulating cholesterol transport in astrocytes. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. 3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiaochun; Lu, Feiran; Trinh, Michael N.; Schmiege, Philip; Seemann, Joachim; Wang, Jiawei; Blobel, Günter

    2017-08-07

    Niemann–Pick C1 (NPC1) and NPC2 proteins are indispensable for the export of LDL-derived cholesterol from late endosomes. Mutations in these proteins result in Niemann–Pick type C disease, a lysosomal storage disease. Despite recent reports of the NPC1 structure depicting its overall architecture, the function of its C-terminal luminal domain (CTD) remains poorly understood even though 45% of NPC disease-causing mutations are in this domain. Here, we report a crystal structure at 3.3 Å resolution of NPC1* (residues 314–1,278), which—in contrast to previous lower resolution structures—features the entire CTD well resolved. Notably, all eight cysteines of the CTD form four disulfide bonds, one of which (C909–C914) enforces a specific loop that in turn mediates an interaction with a loop of the N-terminal domain (NTD). Importantly, this loop and its interaction with the NTD were not observed in any previous structures due to the lower resolution. Our mutagenesis experiments highlight the physiological relevance of the CTD–NTD interaction, which might function to keep the NTD in the proper orientation for receiving cholesterol from NPC2. Additionally, this structure allows us to more precisely map all of the disease-causing mutations, allowing future molecular insights into the pathogenesis of NPC disease.

  15. Nuclear receptors in control of cholesterol transport

    NARCIS (Netherlands)

    van der Veen, Jelske Nynke

    2007-01-01

    Cholesterol is een structurele component van celmembranen en een grondstof voor de aanmaak van steroïde hormonen en galzouten en vervult dus een aantal essentiële fysiologische functies. Een goede balans van cholesterol opname, synthese, afbraak en uitscheiding is noodzakelijk, omdat verhoogde

  16. Cholesterol-Lowering Activity of Tartary Buckwheat Protein.

    Science.gov (United States)

    Zhang, Chengnan; Zhang, Rui; Li, Yuk Man; Liang, Ning; Zhao, Yimin; Zhu, Hanyue; He, Zouyan; Liu, Jianhui; Hao, Wangjun; Jiao, Rui; Ma, Ka Ying; Chen, Zhen-Yu

    2017-03-08

    Previous research has shown that Tartary buckwheat flour is capable of reducing plasma cholesterol. The present study was to examine the effect of rutin and Tartary buckwheat protein on plasma total cholesterol (TC) in hypercholesterolemia hamsters. In the first animal experiment, 40 male hamsters were divided into four groups fed either the control diet or one of the three experimental diets containing 8.2 mmol rutin, 8.2 mmol quercetin, or 2.5 g kg -1 cholestyramine, respectively. Results showed that only cholestyramine but not rutin and its aglycone quercetin decreased plasma TC, which suggested that rutin was not the active ingredient responsible for plasma TC-lowering activity of Tartary buckwheat flour. In the second animal experiment, 45 male hamsters were divided into five groups fed either the control diet or one of the four experimental diets containing 24% Tartary buckwheat protein, 24% rice protein, 24% wheat protein, or 5 g kg -1 cholestyramine, respectively. Tartary buckwheat protein reduced plasma TC more effectively than cholestyramine (45% versus 37%), while rice and wheat proteins only reduced plasma TC by 10-13%. Tartary buckwheat protein caused 108% increase in the fecal excretion of total neutral sterols and 263% increase in the fecal excretion of total acidic sterols. real-time polymerase chain reaction and Western blotting analyses showed that Tartary buckwheat protein affected the gene expression of intestinal Niemann-Pick C1-like protein 1 (NPC1L1), acyl CoA:cholesterol acyltransferase 2 (ACAT2), and ATP binding cassette transporters 5 and 8 (ABCG5/8) in a down trend, whereas it increased the gene expression of hepatic cholesterol-7α -hydroxylase (CYP7A1). It was concluded that Tartary buckwheat protein was at least one of the active ingredients in Tartary buckwheat flour to lower plasma TC, mainly mediated by enhancing the excretion of bile acids via up-regulation of hepatic CYP7A1 and also by inhibiting the absorption of dietary

  17. Comparative Structures and Evolution of Vertebrate Carboxyl Ester Lipase (CEL Genes and Proteins with a Major Role in Reverse Cholesterol Transport

    Directory of Open Access Journals (Sweden)

    Roger S. Holmes

    2011-01-01

    Full Text Available Bile-salt activated carboxylic ester lipase (CEL is a major triglyceride, cholesterol ester and vitamin ester hydrolytic enzyme contained within pancreatic and lactating mammary gland secretions. Bioinformatic methods were used to predict the amino acid sequences, secondary and tertiary structures and gene locations for CEL genes, and encoded proteins using data from several vertebrate genome projects. A proline-rich and O-glycosylated 11-amino acid C-terminal repeat sequence (VNTR previously reported for human and other higher primate CEL proteins was also observed for other eutherian mammalian CEL sequences examined. In contrast, opossum CEL contained a single C-terminal copy of this sequence whereas CEL proteins from platypus, chicken, lizard, frog and several fish species lacked the VNTR sequence. Vertebrate CEL genes contained 11 coding exons. Evidence is presented for tandem duplicated CEL genes for the zebrafish genome. Vertebrate CEL protein subunits shared 53–97% sequence identities; demonstrated sequence alignments and identities for key CEL amino acid residues; and conservation of predicted secondary and tertiary structures with those previously reported for human CEL. Phylogenetic analyses demonstrated the relationships and potential evolutionary origins of the vertebrate CEL family of genes which were related to a nematode carboxylesterase (CES gene and five mammalian CES gene families.

  18. Modulation of cholesterol transport by maternal hypercholesterolemia in human full-term placenta.

    Directory of Open Access Journals (Sweden)

    Ran Zhang

    Full Text Available The significance of maternal cholesterol transporting to the fetus under normal as well as pathological circumstances is less understood. The objective of this study was to observe the effects of maternal hypercholesterolemia on placental cholesterol transportation. Human full-time placenta, maternal and venous cord blood were sampled at delivery from the pregnant women with serum total cholesterol (TC concentrations at third trimester higher than 7.25 mM (n = 19 and the pregnant women with normal TC concentrations (n = 19. Serum lipids and expression of genes related to cholesterol transportation were measured by western blot or real-time PCR. The results indicated that serum TC, high density lipoprotein cholesterol (HDL-C, and low density lipoprotein cholesterol (LDL-C levels were significantly increased, in pregnancies, but decreased in cord blood in hypercholesterolemic group compared to the matched control group. All the subjects were no-drinking, non-smoker, and gestational disease free. The mRNA expression of lipoprotein receptors, including LDLR and VLDLR were significantly increased, while the protein expression of PCSK9 was significantly increased in hypercholesterolemic placenta. In conclusion, maternal hypercholesterolemia might decrease the transportation of cholesterol from mother to fetus because of the high levels of PCSK9 protein expression.

  19. Micellar lipid composition profoundly affects LXR-dependent cholesterol transport across CaCo2 cells

    NARCIS (Netherlands)

    Petruzzelli, Michele; Groen, Albert K.; van Erpecum, Karel J.; Vrins, Carlos; van der Velde, Astrid E.; Portincasa, Piero; Palasciano, Giuseppe; van Berge Henegouwen, Gerard P.; Sasso, Giuseppe Lo; Morgano, Annalisa; Moschetta, Antonio

    2009-01-01

    Intraluminal phospholipids affect micellar solubilization and absorption of cholesterol. We here study cholesterol transport from taurocholate-phospholipid-cholesterol micelles to CaCo2 cells, and associated effects on ABC-A1 mediated cholesterol efflux. Micellar incorporation of

  20. Macrophage mitochondrial damage from StAR transport of 7-hydroperoxycholesterol: implications for oxidative stress-impaired reverse cholesterol transport.

    Science.gov (United States)

    Korytowski, Witold; Wawak, Katarzyna; Pabisz, Pawel; Schmitt, Jared C; Girotti, Albert W

    2014-01-03

    StAR family proteins in vascular macrophages participate in reverse cholesterol transport (RCT). We hypothesize that under pathophysiological oxidative stress, StARs will transport not only cholesterol to macrophage mitochondria, but also pro-oxidant cholesterol hydroperoxides (7-OOHs), thereby impairing early-stage RCT. Upon stimulation with dibutyryl-cAMP, RAW264.7 macrophages exhibited a strong time-dependent induction of mitochondrial StarD1 and plasma membrane ABCA1, which exports cholesterol. 7α-OOH uptake by stimulated RAW cell mitochondria (like cholesterol uptake) was strongly reduced by StarD1 knockdown, consistent with StarD1 involvement. Upon uptake by mitochondria, 7α-OOH (but not redox-inactive 7α-OH) triggered lipid peroxidation and membrane depolarization while reducing ABCA1 upregulation. These findings provide strong initial support for our hypothesis. Copyright © 2013. Published by Elsevier B.V.

  1. Modulating cancer cell survival by targeting intracellular cholesterol transport.

    Science.gov (United States)

    Kuzu, Omer F; Gowda, Raghavendra; Noory, Mohammad A; Robertson, Gavin P

    2017-08-08

    Demand for cholesterol is high in certain cancers making them potentially sensitive to therapeutic strategies targeting cellular cholesterol homoeostasis. A potential approach involves disruption of intracellular cholesterol transport, which occurs in Niemann-Pick disease as a result of acid sphingomyelinase (ASM) deficiency. Hence, a class of lysosomotropic compounds that were identified as functional ASM inhibitors (FIASMAs) might exhibit chemotherapeutic activity by disrupting cancer cell cholesterol homoeostasis. Here, the chemotherapeutic utility of ASM inhibition was investigated. The effect of FIASMAs on intracellular cholesterol levels, cholesterol homoeostasis, cellular endocytosis and signalling cascades were investigated. The in vivo efficacy of ASM inhibition was demonstrated using melanoma xenografts and a nanoparticle formulation was developed to overcome dose-limiting CNS-associated side effects of certain FIASMAs. Functional ASM inhibitors inhibited intracellular cholesterol transport leading to disruption of autophagic flux, cellular endocytosis and receptor tyrosine kinase signalling. Consequently, major oncogenic signalling cascades on which cancer cells were reliant for survival were inhibited. Two tested ASM inhibitors, perphenazine and fluphenazine that are also clinically used as antipsychotics, were effective in inhibiting xenografted tumour growth. Nanoliposomal encapsulation of the perphenazine enhanced its chemotherapeutic efficacy while decreasing CNS-associated side effects. This study suggests that disruption of intracellular cholesterol transport by targeting ASM could be utilised as a potential chemotherapeutic approach for treating cancer.

  2. Heat shock protein 27-derived atheroprotection involves reverse cholesterol transport that is dependent on GM-CSF to maintain ABCA1 and ABCG1 expression in ApoE-/- mice.

    Science.gov (United States)

    Pulakazhi Venu, Vivek Krishna; Adijiang, Ayinuer; Seibert, Tara; Chen, Yong-Xiang; Shi, Chunhua; Batulan, Zarah; O'Brien, Edward R

    2017-06-01

    Recently, we demonstrated that heat shock protein (HSP)-27 is protective against the development of experimental atherosclerosis, reducing plaque cholesterol content by more than 30%. Moreover, elevated HSP-27 levels are predictive of relative freedom from clinical cardiovascular events. HSP-27 signaling occurs via the activation of NF-κB, which induces a marked up-regulation in expression of granulocyte-monocyte colony-stimulating factor (GM-CSF), a cytokine that is known to alter ABC transporters involved in reverse cholesterol transport (RCT). Therefore, we hypothesized that HSP-27-derived GM-CSF has a potent role in impeding plaque formation by promoting macrophage RCT and sought to better characterize this pathway. Treatment of THP-1 cells, RAW-Blue cells, and primary macrophages with recombinant HSP-27 resulted in NF-κB activation via TLR-4 and was inhibited by various pharmacologic blockers of this pathway. Moreover, HSP-27-induced upregulation of GM-CSF expression was dependent on TLR-4 signaling. Recombinant (r)HSP-27 treatment of ApoE -/- female (but not male) mice for 4 wk yielded reductions in plaque area and cholesterol clefts of 33 and 47%, respectively, with no effect on GM-CSF -/- ApoE -/- mice. With 12 wk of rHSP-27 treatment, both female and male mice showed reductions in plaque burden (55 and 42%, respectively) and a 60% reduction in necrotic core area but no treatment effect in GM-CSF -/- ApoE -/- mice. In vitro functional studies revealed that HSP-27 enhanced the expression of ABCA1 and ABCG1, as well as facilitated cholesterol efflux in vitro by ∼10%. These novel findings establish a paradigm for HSP-27-mediated RCT and set the stage for the development of HSP-27 atheroprotective therapeutics.-Pulakazhi Venu, V. K., Adijiang, A., Seibert, T., Chen, Y.-X., Shi, C., Batulan, Z., O'Brien, E. R. Heat shock protein 27-derived atheroprotection involves reverse cholesterol transport that is dependent on GM-CSF to maintain ABCA1 and ABCG1

  3. Alcohol consumption stimulates early steps in reverse cholesterol transport.

    Science.gov (United States)

    van der Gaag, M S; van Tol, A; Vermunt, S H; Scheek, L M; Schaafsma, G; Hendriks, H F

    2001-12-01

    Alcohol consumption is associated with increased HDL cholesterol levels, which may indicate stimulated reverse cholesterol transport. The mechanism is, however, not known. The aim of this study was to evaluate the effects of alcohol consumption on the first two steps of the reverse cholesterol pathway: cellular cholesterol efflux and plasma cholesterol esterification. Eleven healthy middle-aged men consumed four glasses (40 g of alcohol) of red wine, beer, spirits (Dutch gin), or carbonated mineral water (control) daily with evening dinner, for 3 weeks, according to a 4 x 4 Latin square design. After 3 weeks of alcohol consumption the plasma ex vivo cholesterol efflux capacity, measured with Fu5AH cells, was raised by 6.2% (P alcoholic beverages. Plasma cholesterol esterification was increased by 10.8% after alcohol (P = 0.008). Changes were statistically significant after beer and spirits, but not after red wine consumption (P = 0.16). HDL lipids changed after alcohol consumption; HDL total cholesterol, HDL cholesteryl ester, HDL free cholesterol, HDL phospholipids and plasma apolipoprotein A-I all increased (P alcohol consumption stimulates cellular cholesterol efflux and its esterification in plasma. These effects were mostly independent of the kind of alcoholic beverage

  4. Cholesterol 27-hydroxylase but not apolipoprotein apoE contributes to A2A adenosine receptor stimulated reverse cholesterol transport.

    Science.gov (United States)

    Bingham, Taiese Crystal; Parathath, Saj; Tian, Heather; Reiss, Allison; Chan, Edwin; Fisher, Edward A; Cronstein, Bruce N

    2012-02-01

    Movement of free cholesterol between the cellular compartment and acceptor is governed by cholesterol gradients that are determined by several enzymes and reverse cholesterol transport proteins. We have previously demonstrated that adenosine A(2A) receptors inhibit foam cell formation and stimulate production of cholesterol 27-hydroxylase (CYP27A1), an enzyme involved in the conversion of cholesterol to oxysterols. We therefore asked whether the effect of adenosine A(2A) receptors on foam cell formation in vitro is mediated by CYP27A1 or apoE, a carrier for cholesterol in the serum. We found that specific lentiviral siRNA infection markedly reduced apoE or 27-hydroxylase mRNA in THP-1 cells. Despite diminished apoE expression (p CYP27A1 KD cells (4 ± 2%; pCYP27A1 KD cells but reduced efflux in apoE KD cells. These results demonstrate that adenosine A(2A) receptor occupancy diminishes foam cell formation by increasing expression and function of CYP27A1.

  5. Developments in intestinal cholesterol transport and triglyceride absorption.

    Science.gov (United States)

    Paalvast, Yared; de Boer, Jan Freark; Groen, Albert K

    2017-06-01

    To discuss recent advances in research focused on intestinal lipid handling. An important strategy in reducing atherosclerosis and risk of cardiovascular events is to increase the rate of reverse cholesterol transport, including its final step; cholesterol excretion from the body. The rate of removal is determined by a complex interplay between the factors involved in regulation of intestinal cholesterol absorption. One of these factors is a process known as transintestinal cholesterol excretion. This pathway comprises transport of cholesterol directly from the blood, through the enterocyte, into the intestinal lumen. In humans, this pathway accounts for 35% of cholesterol excretion in the feces. Mechanistic studies in mice revealed that, activation of the bile acid receptor farnesoid X receptor increases cholesterol removal via the transintestinal cholesterol excretion pathway as well as decreases plasma cholesterol and triglyceride providing an interesting target for treatment of dyslipidemia in humans. The physical chemical properties of bile acids are under control of farnesoid X receptor and determine intestinal cholesterol and triglyceride solubilization as well as absorption, providing a direct link between these two important factors in the pathogenesis of cardiovascular disease. Besides bile acids, intestinal phospholipids are important for luminal lipid solubilization. Interestingly, phospholipid remodeling through LPCAT3 was shown to be pivotal for uptake of fatty acids by enterocytes, which may provide a mechanistic handle for therapeutic intervention. The importance of the intestine in control of cholesterol and triglyceride homeostasis is increasingly recognized. Recently, novel factors involved in regulation of cholesterol excretion and intestinal triglyceride and fatty acid uptake have been reported and are discussed in this short review.

  6. Potential of BODIPY-cholesterol for analysis of cholesterol transport and diffusion in living cells

    DEFF Research Database (Denmark)

    Wüstner, Daniel; Lund, Frederik Wendelboe; Röhrl, Clemens

    2016-01-01

    is to use intrinsically fluorescent sterols, as dehydroergosterol (DHE), having minimal chemical alteration compared to cholesterol but giving low fluorescence signals in the UV region of the spectrum. Alternatively, one can use dye-tagged cholesterol analogs and in particular BODIPY-cholesterol (BChol...... photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP). We also describe pulse-chase studies from the PM using BChol in direct comparison to DHE. Based on the gathered imaging data, we present a two-step kinetic model for sterol transport between PM and recycling endosomes. In addition, we...

  7. Fatty acids modulate the expression levels of key proteins for cholesterol absorption in Caco-2 monolayer.

    Science.gov (United States)

    Yang, Fang; Chen, Guoxun; Ma, Meihu; Qiu, Ning; Zhu, Lingjiao; Li, Jing

    2018-02-20

    Fatty acids have been shown to modulate intestinal cholesterol absorption in cells and animals, a process that is mediated by several transporter proteins. Of these proteins, Niemann-Pick C1-Like 1 (NPC1L1) is a major contributor to this process. The current study investigates the unknown mechanism by which fatty acids modulate cholesterol absorption. We evaluated the effects of six fatty acids palmitic acid (PAM), oleic acid (OLA), linoleic acid (LNA), arachidonic acid (ARA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on cholesterol uptake and transport in human enterocytes Caco-2 cells, and on the mRNA expression levels of NPC1L1, others proteins (ABCG5, ABCG8, ABCA1, ACAT2, MTP, Caveolin 1, Annexin-2) involved in cholesterol absorption, and SREBP-1 and SREBP-2 that are responsible for lipid metabolism. The polyunsaturated fatty acids (PUFAs), especially for EPA and DHA, dose-dependently inhibited cholesterol uptake and transport in Caco-2 monolayer, while saturated fatty acids (SFAs) and monounsaturated fatty acids (MUFAs) had no inhibitory effects. EPA and DHA inhibited cholesterol absorption in Caco-2 monolayer might be caused by down-regulating NPC1L1 mRNA and protein levels, which were associated with inhibition of SREBP-1/- 2 mRNA expression levels. Results from this study indicate that functional food containing high PUFAs may have potential therapeutic benefit to reduce cholesterol absorption. Further studies on this topic may provide approaches to control lipid metabolism and to promote health.

  8. Transport of cholesterol autoxidation products in rabbit lipoproteins

    International Nuclear Information System (INIS)

    Peng, Shi-Kaung; Phillips, G.A.; Xia, Guang-Zhi; Morin, R.J.

    1987-01-01

    Radiolabeled pure [4- 14 C] cholesterol was kept at 60 0 C under air to autoxidize for 5 weeks, after which approximately 12% cholesterol oxidation products were formed. The mixture, suspended in gelatin, was given to rabbits by gastric gavage. Rabbits were killed 4, 24 and 48 h after treatment. Cholesterol and its autoxidation products were separated by thin-layer chromatography into 5 fractions and radioactivities of each fraction were measured. Percentages of each fraction of cholesterol oxidation products and cholesterol in the original mixture before administration and in the rabbit sera after administration were similar, suggesting that the rates of absorption of cholesterol oxidation products are not significantly different from that of cholesterol. Lipoproteins were fractioned by ultracentrifugation into VLDL, LDL and HDL. Radioactivities of each fraction in lipoproteins separated by thin layer chromatography showed that fractions containing cholestane-3β, 5α, 6β-triol, 7α- and 7β-hydroxycholesterol and 7-ketocholesterol were more selectively transported in VLDL, whereas most of the 25-hydroxycholesterol was present in LDL. HDL contained only minute amounts of cholesterol oxidation products. 22 refs

  9. Functional analysis of candidate ABC transporter proteins for sitosterol transport

    DEFF Research Database (Denmark)

    Albrecht, C; Elliott, J I; Sardini, A

    2002-01-01

    Two ATP-binding cassette (ABC) proteins, ABCG5 and ABCG8, have recently been associated with the accumulation of dietary cholesterol in the sterol storage disease sitosterolemia. These two 'half-transporters' are assumed to dimerize to form the complete sitosterol transporter which reduces the ab...

  10. Functional analysis of candidate ABC transporter proteins for sitosterol transport

    DEFF Research Database (Denmark)

    Albrecht, C; Elliott, J I; Sardini, A

    2002-01-01

    Two ATP-binding cassette (ABC) proteins, ABCG5 and ABCG8, have recently been associated with the accumulation of dietary cholesterol in the sterol storage disease sitosterolemia. These two 'half-transporters' are assumed to dimerize to form the complete sitosterol transporter which reduces...... the absorption of sitosterol and related molecules in the intestine by pumping them back into the lumen. Although mutations altering ABCG5 and ABCG8 are found in affected patients, no functional demonstration of sitosterol transport has been achieved. In this study, we investigated whether other ABC transporters......-specific ABC transporters have acquired specificity to exclude sitosterol and related sterols like cholesterol presumably because the abundance of cholesterol in the membrane would interfere with their action; in consequence, specific transporters have evolved to handle these sterols....

  11. Prenatal Ethanol Exposure Up-Regulates the Cholesterol Transporters ATP-Binding Cassette A1 and G1 and Reduces Cholesterol Levels in the Developing Rat Brain.

    Science.gov (United States)

    Zhou, Chunyan; Chen, Jing; Zhang, Xiaolu; Costa, Lucio G; Guizzetti, Marina

    2014-11-01

    Cholesterol plays a pivotal role in many aspects of brain development; reduced cholesterol levels during brain development, as a consequence of genetic defects in cholesterol biosynthesis, leads to severe brain damage, including microcephaly and mental retardation, both of which are also hallmarks of the fetal alcohol syndrome. We had previously shown that ethanol up-regulates the levels of two cholesterol transporters, ABCA1 (ATP binding cassette-A1) and ABCG1, leading to increased cholesterol efflux and decreased cholesterol content in astrocytes in vitro. In the present study we investigated whether similar effects could be seen in vivo. Pregnant Sprague-Dawley rats were fed liquid diets containing 36% of the calories from ethanol from gestational day (GD) 6 to GD 21. A pair-fed control groups and an ad libitum control group were included in the study. ABCA1 and ABCG1 protein expression and cholesterol and phospholipid levels were measured in the neocortex of female and male fetuses at GD 21. Body weights were decreased in female fetuses as a consequence of ethanol treatments. ABCA1 and ABCG1 protein levels were increased, and cholesterol levels were decreased, in the neocortex of ethanol-exposed female, but not male, fetuses. Levels of phospholipids were unchanged. Control female fetuses fed ad libitum displayed an up-regulation of ABCA1 and a decrease in cholesterol content compared with pair-fed controls, suggesting that a compensatory up-regulation of cholesterol levels may occur during food restriction. Maternal ethanol consumption may affect fetal brain development by increasing cholesterol transporters' expression and reducing brain cholesterol levels. © The Author 2014. Medical Council on Alcohol and Oxford University Press. All rights reserved.

  12. Acrolein impairs the cholesterol transport functions of high density lipoproteins.

    Science.gov (United States)

    Chadwick, Alexandra C; Holme, Rebecca L; Chen, Yiliang; Thomas, Michael J; Sorci-Thomas, Mary G; Silverstein, Roy L; Pritchard, Kirkwood A; Sahoo, Daisy

    2015-01-01

    High density lipoproteins (HDL) are considered athero-protective, primarily due to their role in reverse cholesterol transport, where they transport cholesterol from peripheral tissues to the liver for excretion. The current study was designed to determine the impact of HDL modification by acrolein, a highly reactive aldehyde found in high abundance in cigarette smoke, on the cholesterol transport functions of HDL. HDL was chemically-modified with acrolein and immunoblot and mass spectrometry analyses confirmed apolipoprotein crosslinking, as well as acrolein adducts on apolipoproteins A-I and A-II. The ability of acrolein-modified HDL (acro-HDL) to serve as an acceptor of free cholesterol (FC) from COS-7 cells transiently expressing SR-BI was significantly decreased. Further, in contrast to native HDL, acro-HDL promotes higher neutral lipid accumulation in murine macrophages as judged by Oil Red O staining. The ability of acro-HDL to mediate efficient selective uptake of HDL-cholesteryl esters (CE) into SR-BI-expressing cells was reduced compared to native HDL. Together, the findings from our studies suggest that acrolein modification of HDL produces a dysfunctional particle that may ultimately promote atherogenesis by impairing functions that are critical in the reverse cholesterol transport pathway.

  13. Absorption and transport of cholesterol autoxidation derivatives in rabbits

    International Nuclear Information System (INIS)

    Peng, S.K.; Morin, R.J.; Phillips, G.A.; Xia, G.Z.

    1986-01-01

    Spontaneously autoxidized products of cholesterol have been demonstrated to be angiotoxic and possibly atherogenic. This study investigates the absorption and transport of these cholesterol oxidation derivatives (COD's) as compared to cholesterol. 14 C-labeled cholesterol autoxidized by incubation in a 60 0 C water bath for 5 weeks, then suspended in gelatin and given to New Zealand white rabbits by gastric gavage. Rabbits were sacrificed 24 hours after treatment. COD's were separated by thin layer chromatography (TLC) and radioactivities of each COD and cholesterol were measured. Percentages of each COD and cholesterol in the original mixture before administration and in the rabbits' serum after administration are almost identical, suggesting that the rates of absorption of COD's are not significantly different from that of cholesterol. Lipoproteins were fractionated by ultracentrifugation into VLDL, LDL and HDL. Radioactivities of each COD separated by TLC in each lipoprotein fraction showed that cholestane-3β,5α,6β-triol, 7α- and 7β-hydroxycholesterol and 7-ketocholesterol were predominantly present in VLDL (3 x serum concentration) and 25-hydroxycholesterol was predominantly in LDL (2.5 x serum concentration). HDL contained only minute amounts of COD's. The increased levels of COD's in VLDL and LDL may contribute to the atherogenicity of these lipoprotein

  14. Immuno-histochemical localization of cholesterol binding proteins in ...

    African Journals Online (AJOL)

    Further, cholesterol association in tissue sections was confirmed by using tetramethylrhodamine isothiocyanate (TRITC) labeled florescent antibodies and immuno-blotting of CBPs. Finally, CBPs or cholesterol-carrying proteins were detected intracellularly in midgut epithelial/ microvillus cells named as CBP+. Zymogene ...

  15. Live-cell imaging of new polyene sterols for improved analysis of intracellular cholesterol transport

    DEFF Research Database (Denmark)

    Modzel, M.; Solanko, K. A.; Szomek, M.

    2018-01-01

    Analysis of intracellular cholesterol transport by fluorescence microscopy requires suitable fluorescent analogues of cholesterol. Most existing cholesterol analogues contain lipophilic dyes which can compromise the sterol properties in membranes. An alternative strategy is to introduce additiona...

  16. The cholesterol transporter ABCG1 links cholesterol homeostasis and tumour immunity.

    Science.gov (United States)

    Sag, Duygu; Cekic, Caglar; Wu, Runpei; Linden, Joel; Hedrick, Catherine C

    2015-02-27

    ATP-binding cassette transporter G1 (ABCG1) promotes cholesterol efflux from cells and regulates intracellular cholesterol homeostasis. Here we demonstrate a role of ABCG1 as a mediator of tumour immunity. Abcg1(-/-) mice have dramatically suppressed subcutaneous MB49-bladder carcinoma and B16-melanoma growth and prolonged survival. We show that reduced tumour growth in Abcg1(-/-) mice is myeloid cell intrinsic and is associated with a phenotypic shift of the macrophages from a tumour-promoting M2 to a tumour-fighting M1 within the tumour. Abcg1(-/-) macrophages exhibit an intrinsic bias towards M1 polarization with increased NF-κB activation and direct cytotoxicity for tumour cells in vitro. Overall, our study demonstrates that the absence of ABCG1 inhibits tumour growth through modulation of macrophage function within the tumour, and illustrates a link between cholesterol homeostasis and cancer.

  17. Transporters for the Intestinal Absorption of Cholesterol, Vitamin E, and Vitamin K.

    Science.gov (United States)

    Yamanashi, Yoshihide; Takada, Tappei; Kurauchi, Ryoya; Tanaka, Yusuke; Komine, Toko; Suzuki, Hiroshi

    2017-04-03

    Humans cannot synthesize fat-soluble vitamins such as vitamin E and vitamin K. For this reason, they must be obtained from the diet via intestinal absorption. As the deficiency or excess of these vitamins has been reported to cause several types of diseases and disorders in humans, the intestinal absorption of these nutrients must be properly regulated to ensure good health. However, the mechanism of their intestinal absorption remains poorly understood. Recent studies on cholesterol using genome-edited mice, genome-wide association approaches, gene mutation analyses, and the development of cholesterol absorption inhibitors have revealed that several membrane proteins play crucial roles in the intestinal absorption of cholesterol. Surprisingly, detailed analyses of these cholesterol transporters have revealed that they can also transport vitamin E and vitamin K, providing clues to uncover the molecular mechanisms underlying the intestinal absorption of these fat-soluble vitamins. In this review, we focus on the membrane proteins (Niemann-Pick C1 like 1, scavenger receptor class B type I, cluster of differentiation 36, and ATP-binding cassette transporter A1) that are (potentially) involved in the intestinal absorption of cholesterol, vitamin E, and vitamin K and discuss their physiological and pharmacological importance. We also discuss the related uncertainties that need to be explored in future studies.

  18. Blackcurrant anthocyanins stimulated cholesterol transport via post-transcriptional induction of LDL receptor in Caco-2 cells.

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    Kim, Bohkyung; Bae, Minkyung; Park, Young-Ki; Ma, Hang; Yuan, Tao; Seeram, Navindra P; Lee, Ji-Young

    2018-02-01

    We previously showed that polyphenol-rich blackcurrant extract (BCE) showed a hypocholesterolemic effect in mice fed a high fat diet. As direct cholesterol removal from the body via the intestine has been recently appreciated, we investigated the effect of BCE on the modulation of genes involved in intestinal cholesterol transport using Caco-2 cells as an in vitro model. Caco-2 cells were treated with BCE to determine its effects on mRNA and protein expression of genes important for intestinal cholesterol transport, low-density lipoprotein (LDL) uptake, cellular cholesterol content, and cholesterol transport from basolateral to apical membrane of Caco-2 cell monolayers. Cells were also treated with anthocyanin-rich or -poor fraction of BCE to determine the role of anthocyanin on BCE effects. BCE significantly increased protein levels of LDL receptor (LDLR) without altering its mRNA, which consequently increased LDL uptake into Caco-2 cells. This post-transcriptional induction of LDLR by BCE was markedly attenuated in the presence of rapamycin, an inhibitor of mechanistic target of rapamycin complex 1 (mTORC1). In addition, BCE altered genes involved in cholesterol transport in the enterocytes, including apical and basolateral cholesterol transporters, in such a way that could enhance cholesterol flux from the basolateral to apical side of the enterocytes. Indeed, BCE significantly increased the flux of LDL-derived cholesterol from the basolateral to the apical chamber of Caco-2 monolayer. LDLR protein levels were markedly increased by anthocyanin-rich fraction, but not by anthocyanin-free fraction. mTORC1-dependent post-transcriptional induction of LDLR by BCE anthocyanins drove the transport of LDL-derived cholesterol to the apical side of the enterocytes. This may represent a potential mechanism for the hypocholesterolemic effect of BCE.

  19. Cholesterol-induced conformational changes in the sterol-sensing domain of the Scap protein suggest feedback mechanism to control cholesterol synthesis.

    Science.gov (United States)

    Gao, Yansong; Zhou, Yulian; Goldstein, Joseph L; Brown, Michael S; Radhakrishnan, Arun

    2017-05-26

    Scap is a polytopic protein of endoplasmic reticulum (ER) membranes that transports sterol regulatory element-binding proteins to the Golgi complex for proteolytic activation. Cholesterol accumulation in ER membranes prevents Scap transport and decreases cholesterol synthesis. Previously, we provided evidence that cholesterol inhibition is initiated when cholesterol binds to loop 1 of Scap, which projects into the ER lumen. Within cells, this binding causes loop 1 to dissociate from loop 7, another luminal Scap loop. However, we have been unable to demonstrate this dissociation when we added cholesterol to isolated complexes of loops 1 and 7. We therefore speculated that the dissociation requires a conformational change in the intervening polytopic sequence separating loops 1 and 7. Here we demonstrate such a change using a protease protection assay in sealed membrane vesicles. In the absence of cholesterol, trypsin or proteinase K cleaved cytosolic loop 4, generating a protected fragment that we visualized with a monoclonal antibody against loop 1. When cholesterol was added to these membranes, cleavage in loop 4 was abolished. Because loop 4 is part of the so-called sterol-sensing domain separating loops 1 and 7, these results support the hypothesis that cholesterol binding to loop 1 alters the conformation of the sterol-sensing domain. They also suggest that this conformational change helps transmit the cholesterol signal from loop 1 to loop 7, thereby allowing separation of the loops and facilitating the feedback inhibition of cholesterol synthesis. These insights suggest a new structural model for cholesterol-mediated regulation of Scap activity. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Human immunodeficiency virus impairs reverse cholesterol transport from macrophages.

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    Zahedi Mujawar

    2006-10-01

    Full Text Available Several steps of HIV-1 replication critically depend on cholesterol. HIV infection is associated with profound changes in lipid and lipoprotein metabolism and an increased risk of coronary artery disease. Whereas numerous studies have investigated the role of anti-HIV drugs in lipodystrophy and dyslipidemia, the effects of HIV infection on cellular cholesterol metabolism remain uncharacterized. Here, we demonstrate that HIV-1 impairs ATP-binding cassette transporter A1 (ABCA1-dependent cholesterol efflux from human macrophages, a condition previously shown to be highly atherogenic. In HIV-1-infected cells, this effect was mediated by Nef. Transfection of murine macrophages with Nef impaired cholesterol efflux from these cells. At least two mechanisms were found to be responsible for this phenomenon: first, HIV infection and transfection with Nef induced post-transcriptional down-regulation of ABCA1; and second, Nef caused redistribution of ABCA1 to the plasma membrane and inhibited internalization of apolipoprotein A-I. Binding of Nef to ABCA1 was required for down-regulation and redistribution of ABCA1. HIV-infected and Nef-transfected macrophages accumulated substantial amounts of lipids, thus resembling foam cells. The contribution of HIV-infected macrophages to the pathogenesis of atherosclerosis was supported by the presence of HIV-positive foam cells in atherosclerotic plaques of HIV-infected patients. Stimulation of cholesterol efflux from macrophages significantly reduced infectivity of the virions produced by these cells, and this effect correlated with a decreased amount of virion-associated cholesterol, suggesting that impairment of cholesterol efflux is essential to ensure proper cholesterol content in nascent HIV particles. These results reveal a previously unrecognized dysregulation of intracellular lipid metabolism in HIV-infected macrophages and identify Nef and ABCA1 as the key players responsible for this effect. Our findings

  1. Fish oil promotes macrophage reverse cholesterol transport in mice.

    Science.gov (United States)

    Nishimoto, Tomoyuki; Pellizzon, Michael A; Aihara, Masakazu; Stylianou, Ioannis M; Billheimer, Jeffery T; Rothblat, George; Rader, Daniel J

    2009-10-01

    Fish oil (FO), and specifically omega 3 fatty acids, has favorable effects on cardiovascular outcomes. The aim of this study was to investigate the effects of FO on the process of macrophage reverse cholesterol transport (RCT) in an in vivo mouse model. C57BL/6J mice were fed a FO diet, whereas control mice were fed diets containing alternative sources of fats, soybean oil (SO), and coconut oil (CO) for 4 weeks. Macrophage RCT was assessed by injecting [(3)H]cholesterol-labeled J774 macrophages intraperitoneally into mice. After 48 hours, tissues were harvested and feces were collected. An increase in the excretion of macrophage-derived [(3)H]-tracer recovered in fecal neutral sterols for FO-fed mice was observed (273% versus SO and 182% versus CO). FO also decreased [(3)H]-tracer in hepatic cholesteryl ester compared to SO and CO by 76% and 56%, respectively. To specifically determine the effect of FO on the fate of HDL-derived cholesterol, mice fed FO or SO diets were injected with HDL labeled with [(3)H]cholesteryl oleate, and the disappearance of [(3)H]-tracer from blood and its excretion in feces was measured. There was no significant difference in the fractional catabolic rate of [(3)H]cholesteryl oleate-HDL between the 2 groups. However, there was a 242% increase in the excretion of HDL-derived [(3)H]-tracer recovered in fecal neutral sterols in FO-fed mice, concordant with significantly increased expression of hepatic Abcg5 and Abcg8 mRNA. As measured by this tracer-based assay, FO promoted reverse cholesterol transport, primarily by enhancement of the hepatic excretion of macrophage-derived and HDL-derived cholesterol.

  2. Cholesterol metabolism, transport, and hepatic regulation in dairy cows during transition and early lactation.

    Science.gov (United States)

    Kessler, E C; Gross, J J; Bruckmaier, R M; Albrecht, C

    2014-09-01

    The transition from the nonlactating to the lactating state represents a critical period for dairy cow lipid metabolism because body reserves have to be mobilized to meet the increasing energy requirements for the initiation of milk production. The purpose of this study was to provide a comprehensive overview on cholesterol homeostasis in transition dairy cows by assessing in parallel plasma, milk, and hepatic tissue for key factors of cholesterol metabolism, transport, and regulation. Blood samples and liver biopsies were taken from 50 multiparous Holstein dairy cows in wk 3 antepartum (a.p.), wk 1 postpartum (p.p.), wk 4 p.p., and wk 14 p.p. Milk sampling was performed in wk 1, 4, and 14 p.p. Blood and milk lipid concentrations [triglycerides (TG), cholesterol, and lipoproteins], enzyme activities (phospholipid transfer protein and lecithin:cholesterol acyltransferase) were analyzed using enzymatic assays. Hepatic gene expression patterns of 3-hydroxy-3-methylglutaryl-coenzyme A (HMGC) synthase 1 (HMGCS1) and HMGC reductase (HMGCR), sterol regulatory element-binding factor (SREBF)-1 and -2, microsomal triglyceride transfer protein (MTTP), ATP-binding cassette transporter (ABC) A1 and ABCG1, liver X receptor (LXR) α and peroxisome proliferator activated receptor (PPAR) α and γ were measured using quantitative RT-PCR. Plasma TG, cholesterol, and lipoprotein concentrations decreased from wk 3 a.p. to a minimum in wk 1 p.p., and then gradually increased until wk 14 p.p. Compared with wk 4 p.p., phospholipid transfer protein activity was increased in wk 1 p.p., whereas lecithin:cholesterol acyltransferase activity was lowest at this period. Total cholesterol concentration and mass, and cholesterol concentration in the milk fat fraction decreased from wk 1 p.p. to wk 4 p.p. Both total and milk fat cholesterol concentration were decreased in wk 4 p.p. compared with wk 1 and 14 p.p. The mRNA abundance of genes involved in cholesterol synthesis (SREBF-2, HMGCS1, and

  3. Transintestinal and Biliary Cholesterol Secretion Both Contribute to Macrophage Reverse Cholesterol Transport in Rats-Brief Report.

    Science.gov (United States)

    de Boer, Jan Freark; Schonewille, Marleen; Dikkers, Arne; Koehorst, Martijn; Havinga, Rick; Kuipers, Folkert; Tietge, Uwe J F; Groen, Albert K

    2017-04-01

    Reverse cholesterol transport comprises efflux of cholesterol from macrophages and its subsequent removal from the body with the feces and thereby protects against formation of atherosclerotic plaques. Because of lack of suitable animal models that allow for evaluation of the respective contributions of biliary cholesterol secretion and transintestinal cholesterol excretion (TICE) to macrophage reverse cholesterol transport under physiological conditions, the relative importance of both pathways in this process has remained controversial. To separate cholesterol traffic via the biliary route from TICE, bile flow was mutually diverted between rats, continuously, for 3 days. Groups of 2 weight-matched rats were designated as a pair, and both rats were equipped with cannulas in the bile duct and duodenum. Bile from rat 1 was diverted to the duodenum of rat 2, whereas bile from rat 2 was rerouted to the duodenum of rat 1. Next, rat 1 was injected with [ 3 H]cholesterol-loaded macrophages. [ 3 H]Cholesterol secreted via the biliary route was consequently diverted to rat 2 and could thus be quantified from the feces of that rat. On the other hand, [ 3 H]cholesterol tracer in the feces of rat 1 reflected macrophage-derived cholesterol excreted via TICE. Using this setup, we found that 63% of the label secreted with the fecal neutral sterols had travelled via the biliary route, whereas 37% was excreted via TICE. TICE and biliary cholesterol secretion contribute to macrophage reverse cholesterol transport in rats. The majority of macrophage-derived cholesterol is however excreted via the hepatobiliary route. © 2017 American Heart Association, Inc.

  4. Thermogenic adipocytes promote HDL turnover and reverse cholesterol transport.

    Science.gov (United States)

    Bartelt, Alexander; John, Clara; Schaltenberg, Nicola; Berbée, Jimmy F P; Worthmann, Anna; Cherradi, M Lisa; Schlein, Christian; Piepenburg, Julia; Boon, Mariëtte R; Rinninger, Franz; Heine, Markus; Toedter, Klaus; Niemeier, Andreas; Nilsson, Stefan K; Fischer, Markus; Wijers, Sander L; van Marken Lichtenbelt, Wouter; Scheja, Ludger; Rensen, Patrick C N; Heeren, Joerg

    2017-04-19

    Brown and beige adipocytes combust nutrients for thermogenesis and through their metabolic activity decrease pro-atherogenic remnant lipoproteins in hyperlipidemic mice. However, whether the activation of thermogenic adipocytes affects the metabolism and anti-atherogenic properties of high-density lipoproteins (HDL) is unknown. Here, we report a reduction in atherosclerosis in response to pharmacological stimulation of thermogenesis linked to increased HDL levels in APOE*3-Leiden.CETP mice. Both cold-induced and pharmacological thermogenic activation enhances HDL remodelling, which is associated with specific lipidomic changes in mouse and human HDL. Furthermore, thermogenic stimulation promotes HDL-cholesterol clearance and increases macrophage-to-faeces reverse cholesterol transport in mice. Mechanistically, we show that intravascular lipolysis by adipocyte lipoprotein lipase and hepatic uptake of HDL by scavenger receptor B-I are the driving forces of HDL-cholesterol disposal in liver. Our findings corroborate the notion that high metabolic activity of thermogenic adipocytes confers atheroprotective properties via increased systemic cholesterol flux through the HDL compartment.

  5. Transintestinal and Biliary Cholesterol Secretion Both Contribute to Macrophage Reverse Cholesterol Transport in RatsBrief Report

    NARCIS (Netherlands)

    Boer, de Jan Freark; Schonewille, Marleen; Dikkers, Arne; Koehorst, Martijn; Havinga, Rick; Kuipers, Folkert; Tietge, Uwe J F; Groen, Albert K

    Objective-Reverse cholesterol transport comprises efflux of cholesterol from macrophages and its subsequent removal from the body with the feces and thereby protects against formation of atherosclerotic plaques. Because of lack of suitable animal models that allow for evaluation of the respective

  6. Lack of P2Y(13) in mice fed a high cholesterol diet results in decreased hepatic cholesterol content, biliary lipid secretion and reverse cholesterol transport

    NARCIS (Netherlands)

    Lichtenstein, Laeticia; Serhan, Nizar; Annema, Wijtske; Combes, Guillaume; Robaye, Bernard; Boeynaems, Jean-Marie; Perret, Bertrand; Tietge, Uwe J. F.; Laffargue, Muriel; Martinez, Laurent O.

    2013-01-01

    Background: The protective effect of HDL is mostly attributed to their metabolic function in reverse cholesterol transport (RCT), a process whereby excess cellular cholesterol is taken up from peripheral cells, processed in HDL particles, and later delivered to the liver for further metabolism and

  7. Extended-Release Niacin/Laropiprant Improves Overall Efficacy of Postprandial Reverse Cholesterol Transport.

    Science.gov (United States)

    El Khoury, Petra; Waldmann, Elisa; Huby, Thierry; Gall, Julie; Couvert, Philippe; Lacorte, Jean-Marc; Chapman, John; Frisdal, Eric; Lesnik, Philippe; Parhofer, Klaus G; Le Goff, Wilfried; Guerin, Maryse

    2016-02-01

    Postprandial atherogenic lipoproteins, characterizing high-risk patients, correlate positively with cardiovascular events. Although the effect of niacin on fasting lipids is well established, its impact on atheroprotective reverse cholesterol transport (RCT) pathway and on functional features of circulating lipoproteins during the postprandial state remains indeterminate. We evaluated RCT pathway during postprandial phase in dyslipidemic patients displaying a low high-density lipoprotein (HDL) cholesterol phenotype. Ten subjects on stable statin therapy received 1 g/20 mg extended-release niacin/laropiprant (ERN/LRPT) for 4 weeks followed by 2 g/40 mg ERN/LRPT for additional 8 weeks. At each experimental period, postprandial hypertriglyceridemia and major steps of RCT, including cholesterol efflux from human macrophages, cholesteryl ester transfer protein-mediated cholesteryl ester transfer, and hepatic HDL-cholesteryl ester selective uptake were evaluated. Equally, the capacity of postprandial HDL particles isolated from patients before and after ERN/LRPT treatment to mediate RCT to feces was evaluated in vivo in human apolipoprotein B/cholesteryl ester transfer protein double transgenic mouse model. Compared with baseline, ERN/LRPT significantly reduced postprandial hypertriglyceridemia (incremental area under the curve-triglyceride: -53%; P=0.02). Postprandial increase in endogenous plasma cholesteryl ester transfer protein activity was completely abolished after ERN/LRPT treatment. Despite a slight reduction in plasma cholesterol efflux capacity from human THP-1 macrophages, evaluation of global RCT efficacy by combining both ex vivo and in vivo approaches indicate that postprandial HDL particles formed under ERN/LRPT therapy displayed a greater capacity for HDL-mediated RCT to feces. ERN/LRPT treatment efficiently attenuates atherogenic postprandial lipemia and stimulates HDL-mediated cholesterol return to the liver and elimination into feces during

  8. Interaction of cholesterol-crystallization-promoting proteins with vesicles

    NARCIS (Netherlands)

    de Bruijn, M. A.; Goldhoorn, B. G.; Zijlstra, A. I.; Tytgat, G. N.; Groen, A. K.

    1995-01-01

    In this study, the interaction of mucin and concanavalin A-binding proteins isolated from human bile with cholesterol/phospholipid vesicles was investigated. Using resonance energy transfer assays originally developed by Struck, Hoekstra and Pagano [(1981) Biochemistry 20, 4093-4099], no significant

  9. Effects of cholesterol transport inhibitor U18666A on APP metabolism in rat primary astrocytes.

    Science.gov (United States)

    Yang, Hongyan; Wang, Yanlin; Kar, Satyabrata

    2017-11-01

    Amyloid β (Aβ) peptides generated from the amyloid precursor protein (APP) play an important role in the degeneration of neurons and development of Alzheimer's disease (AD). Current evidence indicates that high levels of cholesterol-which increase the risk of developing AD-can influence Aβ production in neurons. However, it remains unclear how altered level/subcellular distribution of cholesterol in astrocytes can influence APP metabolism. In this study, we evaluated the effects of cholesterol transport inhibitor U18666A-a class II amphiphile that triggers redistribution of cholesterol within the endosomal-lysosomal (EL) system-on APP levels and metabolism in rat primary cultured astrocytes. Our results revealed that U18666A increased the levels of the APP holoprotein and its cleaved products (α-/β-/η-CTFs) in cultured astrocytes, without altering the total levels of cholesterol or cell viability. The cellular levels of Aβ 1-40 were also found to be markedly increased, while secretory levels of Aβ 1-40 were decreased in U18666A-treated astrocytes. We further report a corresponding increase in the activity of the enzymes regulating APP processing, such as α-secretase, β-secretase, and γ-secretase as a consequence of U18666A treatment. Additionally, APP-cleaved products are partly accumulated in the lysosomes following cholesterol sequestration within EL system possibly due to decreased clearance. Interestingly, serum delipidation attenuated enhanced levels of APP and its cleaved products following U18666A treatment. Collectively, these results suggest that cholesterol sequestration within the EL system in astrocytes can influence APP metabolism and the accumulation of APP-cleaved products including Aβ peptides, which can contribute to the development of AD pathology. © 2017 Wiley Periodicals, Inc.

  10. LDL Receptor-Related Protein-1 (LRP1 Regulates Cholesterol Accumulation in Macrophages.

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    Anna P Lillis

    Full Text Available Within the circulation, cholesterol is transported by lipoprotein particles and is taken up by cells when these particles associate with cellular receptors. In macrophages, excessive lipoprotein particle uptake leads to foam cell formation, which is an early event in the development of atherosclerosis. Currently, mechanisms responsible for foam cell formation are incompletely understood. To date, several macrophage receptors have been identified that contribute to the uptake of modified forms of lipoproteins leading to foam cell formation, but the in vivo contribution of the LDL receptor-related protein 1 (LRP1 to this process is not known [corrected]. To investigate the role of LRP1 in cholesterol accumulation in macrophages, we generated mice with a selective deletion of LRP1 in macrophages on an LDL receptor (LDLR-deficient background (macLRP1-/-. After feeding mice a high fat diet for 11 weeks, peritoneal macrophages isolated from Lrp+/+ mice contained significantly higher levels of total cholesterol than those from macLRP1-/- mice. Further analysis revealed that this was due to increased levels of cholesterol esters. Interestingly, macLRP1-/- mice displayed elevated plasma cholesterol and triglyceride levels resulting from accumulation of large, triglyceride-rich lipoprotein particles in the circulation. This increase did not result from an increase in hepatic VLDL biosynthesis, but rather results from a defect in catabolism of triglyceride-rich lipoprotein particles in macLRP1-/- mice. These studies reveal an important in vivo contribution of macrophage LRP1 to cholesterol homeostasis.

  11. LXR driven induction of HDL-cholesterol is independent of intestinal cholesterol absorption and ABCA1 protein expression.

    Science.gov (United States)

    Kannisto, Kristina; Gåfvels, Mats; Jiang, Zhao-Yan; Slätis, Katharina; Hu, Xiaoli; Jorns, Carl; Steffensen, Knut R; Eggertsen, Gösta

    2014-01-01

    We investigated whether: (1) liver X receptor (LXR)-driven induction of high-density lipoprotein cholesterol (HDL-C) and other LXR-mediated effects on cholesterol metabolism depend on intestinal cholesterol absorption; and (2) combined treatment with the LXR agonist GW3965 and the cholesterol absorption inhibitor ezetimibe results in synergistic effects on cholesterol metabolism that could be beneficial for treatment of atherosclerosis. Mice were fed 0.2 % cholesterol and treated with GW3965+ezetimibe, GW3965 or ezetimibe. GW3965+ezetimibe treatment elevated serum HDL-C and Apolipoprotein (Apo) AI, effectively reduced the intestinal cholesterol absorption and increased the excretion of faecal neutral sterols. No changes in intestinal ATP-binding cassette (ABC) A1 or ABCG5 protein expression were observed, despite increased mRNA expression, while hepatic ABCA1 was slightly reduced. The combined treatment caused a pronounced down-regulation of intestinal Niemann-Pick C1-like 1 (NPC1L1) and reduced hepatic and intestinal cholesterol levels. GW3965 did not affect the intestinal cholesterol absorption, but increased serum HDL-C and ApoAI levels. GW3965 also increased Apoa1 mRNA levels in primary mouse hepatocytes and HEPA1-6 cells. Ezetimibe reduced the intestinal cholesterol absorption, ABCA1 and ABCG5, but did not affect the serum HDL-C or ApoAI levels. Thus, the LXR-driven induction of HDL-C and ApoAI was independent of the intestinal cholesterol absorption and increased expression of intestinal or hepatic ABCA1 was not required. Inhibited influx of cholesterol via NPC1L1 and/or low levels of intracellular cholesterol prevented post-transcriptional expression of intestinal ABCA1 and ABCG5, despite increased mRNA levels. Combined LXR activation and blocked intestinal cholesterol absorption induced effective faecal elimination of cholesterol.

  12. The Role of Omega-3 Fatty Acids in Reverse Cholesterol Transport: A Review

    Directory of Open Access Journals (Sweden)

    Alex Pizzini

    2017-10-01

    Full Text Available The beneficial effects of omega-3 polyunsaturated fatty acids (n-3 PUFAs on cardiovascular disease have been studied extensively. However, it remains unclear to what extent n-3 PUFAs may impact Reverse Cholesterol Transport (RCT. RCT describes a mechanism by which excess cholesterol from peripheral tissues is transported to the liver for hepatobiliary excretion, thereby inhibiting foam cell formation and the development of atherosclerosis. The aim of this review is to summarize the literature and to provide an updated overview of the effects of n-3 PUFAs on key players in RCT, including apoliprotein AI (apoA-I, ATP-binding cassette transporter A1 (ABCA1, ABCG1, apoE, scavenger receptor class B type I (SR-BI, cholesteryl ester transfer protein (CETP, low-density lipoprotein receptor (LDLr, cholesterol 7 alpha-hydroxylase (CYP7A1 and ABCG5/G8. Based on current knowledge, we conclude that n-3 PUFAs may beneficially affect RCT, mainly by influencing high-density lipoprotein (HDL remodeling and by promoting hepatobiliary sterol excretion.

  13. [A history and review of cholesterol ester transfer protein inhibitors and their contribution to the understanding of the physiology and pathophysiology of high density lipoprotein].

    Science.gov (United States)

    Corral, Pablo; Schreier, Laura

    2014-01-01

    There is irrefutable evidence that statins reduce the risk of cardiovascular events in a magnitude proportional to the intensity of the decrease in cholesterol transport by the low density lipoproteins. Despite this great advance there is still a residual risk of cardiovascular events. For this reason, an increase in the levels of high density lipoprotein is considered in order to boost the main action of this lipoprotein, which is reverse cholesterol transport. Distinct classes of evidence (epidemiological, genetic, and pathophysiological) show that the inhibition and/or modulation of cholesterol ester transfer protein increases plasma high density lipoprotein-cholesterol levels. The main reason for presenting this review is to look at the physiology of cholesterol ester transfer protein, its interrelationship with high density lipoproteins, and to give an update on the development of different cholesterol ester transfer protein inhibitor/modulator molecules. Copyright © 2013 Elsevier España, S.L. y SEA. All rights reserved.

  14. Evolutionary origin of the mitochondrial cholesterol transport machinery reveals a universal mechanism of steroid hormone biosynthesis in animals.

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    Jinjiang Fan

    Full Text Available Steroidogenesis begins with the transport of cholesterol from intracellular stores into mitochondria via a series of protein-protein interactions involving cytosolic and mitochondrial proteins located at both the outer and inner mitochondrial membranes. In adrenal glands and gonads, this process is accelerated by hormones, leading to the production of high levels of steroids that control tissue development and function. A hormone-induced multiprotein complex, the transduceosome, was recently identified, and is composed of cytosolic and outer mitochondrial membrane proteins that control the rate of cholesterol entry into the outer mitochondrial membrane. More recent studies unveiled the steroidogenic metabolon, a bioactive, multimeric protein complex that spans the outer-inner mitochondrial membranes and is responsible for hormone-induced import, segregation, targeting, and metabolism of cholesterol by cytochrome P450 family 11 subfamily A polypeptide 1 (CYP11A1 in the inner mitochondrial membrane. The availability of genome information allowed us to systematically explore the evolutionary origin of the proteins involved in the mitochondrial cholesterol transport machinery (transduceosome, steroidogenic metabolon, and signaling proteins, trace the original archetype, and predict their biological functions by molecular phylogenetic and functional divergence analyses, protein homology modeling and molecular docking. Although most members of these complexes have a history of gene duplication and functional divergence during evolution, phylogenomic analysis revealed that all vertebrates have the same functional complex members, suggesting a common mechanism in the first step of steroidogenesis. An archetype of the complex was found in invertebrates. The data presented herein suggest that the cholesterol transport machinery is responsible for steroidogenesis among all vertebrates and is evolutionarily conserved throughout the entire animal kingdom.

  15. Common structural features of cholesterol binding sites in crystallized soluble proteins.

    Science.gov (United States)

    Bukiya, Anna N; Dopico, Alejandro M

    2017-06-01

    Cholesterol-protein interactions are essential for the architectural organization of cell membranes and for lipid metabolism. While cholesterol-sensing motifs in transmembrane proteins have been identified, little is known about cholesterol recognition by soluble proteins. We reviewed the structural characteristics of binding sites for cholesterol and cholesterol sulfate from crystallographic structures available in the Protein Data Bank. This analysis unveiled key features of cholesterol-binding sites that are present in either all or the majority of sites: i ) the cholesterol molecule is generally positioned between protein domains that have an organized secondary structure; ii ) the cholesterol hydroxyl/sulfo group is often partnered by Asn, Gln, and/or Tyr, while the hydrophobic part of cholesterol interacts with Leu, Ile, Val, and/or Phe; iii ) cholesterol hydrogen-bonding partners are often found on α-helices, while amino acids that interact with cholesterol's hydrophobic core have a slight preference for β-strands and secondary structure-lacking protein areas; iv ) the steroid's C21 and C26 constitute the "hot spots" most often seen for steroid-protein hydrophobic interactions; v ) common "cold spots" are C8-C10, C13, and C17, at which contacts with the proteins were not detected. Several common features we identified for soluble protein-steroid interaction appear evolutionarily conserved. Copyright © 2017 by the American Society for Biochemistry and Molecular Biology, Inc.

  16. High-density lipoproteincholesterol, reverse cholesterol transport, and cardiovascular risk: a tale of genetics?

    Directory of Open Access Journals (Sweden)

    Giovanni Cimmino

    2013-10-01

    Full Text Available Cholesterol deposition plays a central role in atherogenesis. The accumulation of lipid material is the result of an imbalance between the influx and efflux of cholesterol within the arterial wall. High levels of plasma low-density lipoprotein-cholesterol are considered the major mechanism responsible for the influx and accumulation of cholesterol in the arterial wall, while high-density lipoprotein (HDL- cholesterol seems responsible for its efflux. The mechanism by which cholesterol is removed from extra-hepatic organs and delivered to the liver for its catabolism and excretion is called reverse cholesterol transport (RCT. Epidemiological evidence has associated high levels of HDL-cholesterol/ApoA-I with protection against atherosclerotic disease, but the ultimate mechanism(s responsible for the beneficial effect is not well established. HDLs are synthesized by the liver and small intestine and released to the circulation as a lipid-poor HDL (nascent HDL, mostly formed by ApoA-I and phospholipids. Through their metabolic maturation, HDLs interact with the ABCA1 receptor in the macrophage surface increasing their lipid content by taking phospholipids and cholesterol from macrophages becoming mature HDL. The cholesterol of the HDLs is transported to the liver, via the scavenger receptor class B, type I, for further metabolization and excretion to the intestines in the form of bile acids and cholesterol, completing the process of RCT. It is clear that an inherited mutation or acquired abnormality in any of the key players in RCT mat affect the atherosclerotic process.

  17. Effects of Cholesterol on the Thermodynamics and Kinetics of Passive Transport of Water through Lipid Membranes.

    Science.gov (United States)

    Issack, Bilkiss B; Peslherbe, Gilles H

    2015-07-23

    While it has long been known that cholesterol reduces the permeability of biological membranes to water, the exact mechanism by which cholesterol influences transmembrane permeation is still unclear. The thermodynamic and kinetic contributions to the transport of water across mixed DPPC/cholesterol bilayers of different composition are thus examined by molecular dynamics simulations. Our analyses show that cholesterol decreases transmembrane permeability to water mainly by altering the thermodynamics of water transport. In particular, the free-energy barrier to permeation is magnified in the dense bilayer interior and the partitioning of water is significantly lowered. The changes are observed to correlate quantitatively well with the cholesterol-dependent density and thickness of the bilayers. In contrast, diffusion coefficients are relatively insensitive to cholesterol concentration, except in the sparsely populated center of the bilayer. Diffusion of water in cholesterol-containing bilayers appears to be related to changes in the free area in the middle of the bilayer and to the solute cross-sectional area in the denser hydrophobic regions. Overall, cholesterol is found to have an inhibitory effect on the permeation of water at all concentrations investigated, although bilayers containing cholesterol concentrations up to 20 mol % display a more dramatic dependence on cholesterol content than at higher concentrations. Our results show that it is possible to quantitatively reproduce the relative effects of cholesterol on lipid bilayer permeability from molecular dynamics simulations.

  18. [Nicotinic acid increases cellular transport of high density lipoprotein cholesterol in patients with hypoalphalipoproteinemia].

    Science.gov (United States)

    Figueroa, Catalina; Droppelmann, Katherine; Quiñones, Verónica; Amigo, Ludwig; Mendoza, Camila; Serrano, Valentina; Véjar, Margarita; Maiz, Alberto; Rigotti, Attilio

    2015-09-01

    Plasma high density lipoproteins (HDL) are involved in reverse cholesterol transport mediated by the scavenger receptor class B type I (SR-BI). Nicotinic acid increases HDL cholesterol levels, even though its specific impact on SR-BI dependent-cellular cholesterol transport remains unknown. To determine the effect of nicotinic acid on HDL particle functionality in cholesterol efflux and uptake mediated by SR-BI in cultured cells in hypoalphalipoproteinemic patients. In a pilot study, eight patients with low HDL (≤ 40 mg/dL) were treated with extended release nicotinic acid. HDL cholesterol and phospholipid levels, HDL2 and HDL3 fractions and HDL particle sizes were measured at baseline and post-therapy. Before and after nicotinic acid treatment, HDL particles were used for cholesterol transport studies in cells transfected with SR-BI. Nicotinic acid treatment raised total HDL cholesterol and phospholipids, HDL2 levels as well as HDL particle size. Nicotinic acid significantly increased HDL cholesterol efflux and uptake capacity mediated by SR-BI in cultured cells. Nicotinic acid therapy increases SR-BI-dependent HDL cholesterol transport in cultured cells, establishing a new cellular mechanism by which this lipid-lowering drug appears to modulate HDL metabolism in patients with hypoalphalipoproteinemia.

  19. Prenatal Ethanol Exposure Up-Regulates the Cholesterol Transporters ATP-Binding Cassette A1 and G1 and Reduces Cholesterol Levels in the Developing Rat Brain

    OpenAIRE

    Zhou, Chunyan; Chen, Jing; Zhang, Xiaolu; Costa, Lucio G.; Guizzetti, Marina

    2014-01-01

    Aims: Cholesterol plays a pivotal role in many aspects of brain development; reduced cholesterol levels during brain development, as a consequence of genetic defects in cholesterol biosynthesis, leads to severe brain damage, including microcephaly and mental retardation, both of which are also hallmarks of the fetal alcohol syndrome. We had previously shown that ethanol up-regulates the levels of two cholesterol transporters, ABCA1 (ATP binding cassette-A1) and ABCG1, leading to increased cho...

  20. Effect of Cholesterol on the Branched-Chain Amino Acid Transport System of Streptococcus cremoris

    NARCIS (Netherlands)

    Zheng, Tan; Driessen, Arnold J.M.; Konings, Wilhelmus

    The effect of cholesterol on the activity of the branched-chain amino acid transport system of Streptococcus cremoris was studied in membrane vesicles of S. cremoris fused with liposomes made of egg yolk phosphatidylcholine, soybean phosphatidylethanolamine, and various amounts of cholesterol.

  1. Impact of LDL apheresis on atheroprotective reverse cholesterol transport pathway in familial hypercholesterolemia

    NARCIS (Netherlands)

    Orsoni, Alexina; Villard, Elise F.; Bruckert, Eric; Robillard, Paul; Carrie, Alain; Bonnefont-Rousselot, Dominique; Chapman, M. John; Dallinga-Thie, Geesje M.; Le Goff, Wilfried; Guerin, Maryse

    2012-01-01

    In familial hypercholesterolemia (FH), low HDL cholesterol (HDL-C) levels are associated with functional alterations of HDL particles that reduce their capacity to mediate the reverse cholesterol transport (RCT) pathway. The objective of this study was to evaluate the consequences of LDL apheresis

  2. The Role of Omega-3 Fatty Acids in Reverse Cholesterol Transport: A Review

    OpenAIRE

    Pizzini, Alex; Lunger, Lukas; Demetz, Egon; Hilbe, Richard; Weiss, Guenter; Ebenbichler, Christoph; Tancevski, Ivan

    2017-01-01

    The beneficial effects of omega-3 polyunsaturated fatty acids (n-3 PUFAs) on cardiovascular disease have been studied extensively. However, it remains unclear to what extent n-3 PUFAs may impact Reverse Cholesterol Transport (RCT). RCT describes a mechanism by which excess cholesterol from peripheral tissues is transported to the liver for hepatobiliary excretion, thereby inhibiting foam cell formation and the development of atherosclerosis. The aim of this review is to summarize the literatu...

  3. Transport of Proteins through Nanopores

    Science.gov (United States)

    Luan, Binquan

    In biological cells, a malfunctioned protein (such as misfolded or damaged) is degraded by a protease in which an unfoldase actively drags the protein into a nanopore-like structure and then a peptidase cuts the linearized protein into small fragments (i.e. a recycling process). Mimicking this biological process, many experimental studies have focused on the transport of proteins through a biological protein pore or a synthetic solid-state nanopore. Potentially, the nanopore-based sensors can provide a platform for interrogating proteins that might be disease-related or be targeted by a new drug molecule. The single-profile of a protein chain inside an extremely small nanopore might even permit the sequencing of the protein. Here, through all-atom molecular dynamics simulations, I will show various types of protein transport through a nanopore and reveal the nanoscale mechanics/energetics that plays an important role governing the protein transport.

  4. Water-transporting proteins

    DEFF Research Database (Denmark)

    Zeuthen, Thomas

    2010-01-01

    . In the K(+)/Cl(-) and the Na(+)/K(+)/2Cl(-) cotransporters, water is entirely cotransported, while water transport in glucose uniporters and Na(+)-coupled transporters of nutrients and neurotransmitters takes place by both osmosis and cotransport. The molecular mechanism behind cotransport of water...... transport. Epithelial water transport is energized by the movements of ions, but how the coupling takes place is uncertain. All epithelia can transport water uphill against an osmotic gradient, which is hard to explain by simple osmosis. Furthermore, genetic removal of aquaporins has not given support...... to osmosis as the exclusive mode of transport. Water cotransport can explain the coupling between ion and water transport, a major fraction of transepithelial water transport and uphill water transport. Aquaporins enhance water transport by utilizing osmotic gradients and cause the osmolarity...

  5. The ABCG5/8 Cholesterol Transporter and Myocardial Infarction Versus Gallstone Disease

    DEFF Research Database (Denmark)

    Stender, Stefan; Frikke-Schmidt, Ruth; Nordestgaard, Børge G

    2014-01-01

    OBJECTIVES: The study sought to test the hypothesis that genetic variation in ABCG5/8, the transporter responsible for intestinal and hepatobiliary cholesterol efflux, may simultaneously influence plasma and biliary cholesterol levels, and hence risk of myocardial infarction (MI) and gallstone...... disease in opposite directions. BACKGROUND: High plasma levels of low-density lipoprotein (LDL) cholesterol are a causal risk factor for MI, whereas high levels of biliary cholesterol promote gallstone formation. METHODS: A total of 60,239 subjects from Copenhagen were included, including 5,647 with MI...... and 3,174 with symptomatic gallstone disease. Subjects were genotyped for 6 common, nonsynonymous and functional variants in ABCG5/8, and a combined weighted genotype score was calculated. RESULTS: Combined, weighted genotype scores were associated with stepwise decreases in LDL cholesterol of up to 5...

  6. Reverse Cholesterol Transport Is Increased in Germ-Free Mice-Brief Report.

    Science.gov (United States)

    Mistry, Rima H; Verkade, Henkjan J; Tietge, Uwe J F

    2017-03-01

    The intestinal microbiota is emerging as a clinically relevant modulator of atherosclerotic risk. Reverse cholesterol transport (RCT) is an atheroprotective metabolic pathway. How the microbiota impacts RCT has not been investigated. Therefore, the aim of this study was to characterize (cholesterol) metabolism and RCT in germ-free mice compared with conventional mice. In chow-fed germ-free mice, plasma cholesterol was unchanged, whereas liver cholesterol content was higher (1.5-fold; P cholesterol (2-fold; P 2-fold ( P 2-fold. Thereby, our results support the importance of intestinal bacteria for metabolic regulation and indicate that specific targeting of the microbiota bears therapeutic potential to prevent and treat cardiovascular disease. © 2017 American Heart Association, Inc.

  7. Omega 3 fatty acids promote macrophage reverse cholesterol transport in hamster fed high fat diet.

    Directory of Open Access Journals (Sweden)

    Fatima Kasbi Chadli

    Full Text Available The aim of this study was to investigate macrophage reverse cholesterol transport (RCT in hamster, a CETP-expressing species, fed omega 3 fatty acids (ω3PUFA supplemented high fat diet (HFD. Three groups of hamsters (n = 6/group were studied for 20 weeks: 1 control diet: Control, 2 HFD group: HF and 3 HFD group supplemented with ω3PUFA (EPA and DHA: HFω3. In vivo macrophage-to-feces RCT was assessed after an intraperitoneal injection of (3H-cholesterol-labelled hamster primary macrophages. Compared to Control, HF presented significant (p<0.05 increase in body weight, plasma TG (p<0.01 and cholesterol (p<0.001 with an increase in VLDL TG and in VLDL and LDL cholesterol (p<0.001. Compared to HF, HFω3 presented significant decrease in body weight. HFω3 showed less plasma TG (p<0.001 and cholesterol (p<0.001 related to a decrease in VLDL TG and HDL cholesterol respectively and higher LCAT activity (p<0.05 compared to HF. HFω3 showed a higher fecal bile acid excretion (p<0.05 compared to Control and HF groups and higher fecal cholesterol excretion (p<0.05 compared to HF. This increase was related to higher gene expression of ABCG5, ABCA1 and SR-B1 in HFω3 compared to Control and HF groups (<0.05 and in ABCG1 and CYP7A1 compared to HF group (p<0.05. A higher plasma efflux capacity was also measured in HFω3 using (3H- cholesterol labeled Fu5AH cells. In conclusion, EPA and DHA supplementation improved macrophage to feces reverse cholesterol transport in hamster fed HFD. This change was related to the higher cholesterol and fecal bile acids excretion and to the activation of major genes involved in RCT.

  8. Immuno-histochemical localization of cholesterol binding proteins in ...

    African Journals Online (AJOL)

    SAM

    2014-07-09

    Jul 9, 2014 ... Positive control tissue sections were stained with Sudan Black-B for microscopic visualization of cholesterol binding sites. Further, cholesterol .... 6 cm) by holes made. In the central well, 100 µL of polyclonal .... equivalence of both substances or formation of a concen- tration gradient (Figures 2a to d).

  9. Cholesterol as a co-solvent and a ligand for membrane proteins

    Science.gov (United States)

    Song, Yuanli; Kenworthy, Anne K; Sanders, Charles R

    2014-01-01

    As of mid 2013 a Medline search on “cholesterol” yielded over 200,000 hits, reflecting the prominence of this lipid in numerous aspects of animal cell biology and physiology under conditions of health and disease. Aberrations in cholesterol homeostasis underlie both a number of rare genetic disorders and contribute to common sporadic and complex disorders including heart disease, stroke, type II diabetes, and Alzheimer's disease. The corresponding author of this review and his lab stumbled only recently into the sprawling area of cholesterol research when they discovered that the amyloid precursor protein (APP) binds cholesterol, a topic covered by the Hans Neurath Award lecture at the 2013 Protein Society Meeting. Here, we first provide a brief overview of cholesterol-protein interactions and then offer our perspective on how and why binding of cholesterol to APP and its C99 domain (β-CTF) promotes the amyloidogenic pathway, which is closely related to the etiology of Alzheimer's disease. PMID:24155031

  10. Inositol transport proteins.

    Science.gov (United States)

    Schneider, Sabine

    2015-04-28

    The cyclic polyol myo-inositol is a key molecule in many different metabolic pathways among all organisms; in addition, it is fundamental for osmotic balance in the mammalian brain. This review sums up inositol transporters from eukaryotic organisms, elucidating their vital role in regulating the intracellular distribution and uptake of inositol. They can be divided into two groups according to their transport mechanisms: (1) sodium ion coupled inositol transporters that belong to the Solute Carrier Families 5 and 6-like Superfamily and, (2) proton coupled inositol symporters that are members of the Major Facilitator Superfamily. Intriguingly members of both families offer promising targets for medical treatment of a variety of diseases. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  11. Antihyperlipidemic effect of sesame (Sesamum indicum L.) protein isolate in rats fed a normal and high cholesterol diet.

    Science.gov (United States)

    Biswas, Arundhati; Dhar, Pubali; Ghosh, Santinath

    2010-01-01

    The dietary influence of sesame protein isolate (protein content 91.5%), produced from dehulled, defatted sesame meal, on blood and tissue lipid profile and lipid peroxidation has been assessed in normal and hypercholesterolemic rats. To evaluate their hypocholesterolemic and antioxidative activity in vivo, we fed 18% sesame protein isolate with or without 2% cholesterol in comparison with casein to rats for 28 d. We determined plasma total protein, total cholesterol, high-density lipoprotein (HDL)-cholesterol, low-density lipoprotein (LDL)-cholesterol, triacylglycerol as well as susceptibility of plasma and erythrocyte membrane lipid to oxidation ex vivo. Liver tissue lipid, cholesterol, phospholipids, and lipid peroxidations were also determined. The total cholesterol, LDL-cholesterol and triacylglycerol levels were significantly reduced in the sesame protein isolate and isolate containing cholesterol group than the corresponding control casein groups. HDL-cholesterol level was also increased in sesame protein isolate (41%) and protein isolate containing cholesterol group (38%) than the corresponding control casein and casein containing cholesterol groups. There was 49% and 64% lowering of plasma lipid peroxidation as well as 36% and 56% lowering of lipoprotein oxidation susceptibility (LOS) in the 2 experimental groups (sesame protein isolate and isolate containing cholesterol group) than the corresponding control (casein and casein containing cholesterol) groups. There was significant lowering of erythrocyte membrane lipid peroxidation (68% and 63% lowering in sesame protein isolate and isolate containing cholesterol groups) and liver lipid peroxidation (61% and 76% lowering in the 2 experimental groups than the corresponding control casein groups). Therefore, our results indicate that sesame protein isolate decreases cholesterol concentration in plasma, increases HDL-cholesterol, and also decreases plasma and erythrocyte membrane lipid peroxidation with or

  12. Effects of ginsenosides on the expression of cytochrome P450s and transporters involved in cholesterol metabolism.

    Science.gov (United States)

    Kawase, Atsushi; Yamada, Ayano; Gamou, Yuko; Tahara, Chika; Takeshita, Fumiaki; Murata, Kazuya; Matsuda, Hideaki; Samukawa, Keiichi; Iwaki, Masahiro

    2014-04-01

    An extract from red ginseng (steamed and dried roots of Panax ginseng C.A. Meyer; RGE) has been shown to promote cholesterol metabolism in the liver. We have reported that RGE induced the hepatic expression of cytochrome P450 (CYP)7A1, involved in cholesterol metabolism. Other cholesterol metabolism-related proteins, such as CYP8B1, CYP27A1, multidrug resistance-associated protein (MRP)2, MRP3, and Na(+) taurocholate cotransporting polypeptide (NTCP), are involved in cholesterol metabolism. The purpose of this study was to clarify whether RGE affected mRNA expression of cholesterol metabolism-related CYPs and transporters in the liver of hypercholesterolemic rats and rat primary hepatocytes. In-vivo studies showed little differences in CYP8B1, CYP27A1, MRP2, MRP3, and NTCP mRNA expression levels between hypercholesterolemic rats with or without RGE treatments. However, the disruption of the membrane localization of MRP2 was suppressed by RGE treatments in hypercholesterolemic rats. In-vitro studies using rat primary hepatocytes showed upregulation of CYP8B1 and MRP2 mRNA by the addition of RGE (100 and 500 μg/mL). We further examined which ginsenosides contributed to the upregulation of CYP8B1 and MRP2 mRNA levels. Ginsenoside Re enhanced the mRNA level of CYP8B1, whereas ginsenosides Rb2 and Rg2 enhanced MRP2 mRNA levels. These results suggest that the in-vitro exposure of hepatocytes to RGE or some ginsenosides could lead to upregulation of CYP8B1 and MRP2, resulting in the alteration of biosynthesis and disposition of bile acids.

  13. Artificial oxygen transport protein

    Science.gov (United States)

    Dutton, P. Leslie

    2014-09-30

    This invention provides heme-containing peptides capable of binding molecular oxygen at room temperature. These compounds may be useful in the absorption of molecular oxygen from molecular oxygen-containing atmospheres. Also included in the invention are methods for treating an oxygen transport deficiency in a mammal.

  14. Genetic variation in ABC transporter A1 contributes to HDL cholesterol in the general population

    DEFF Research Database (Denmark)

    Frikke-Schmidt, Ruth; Nordestgaard, Børge G; Jensen, Gorm B

    2004-01-01

    Homozygosity for mutations in ABC transporter A1 (ABCA1) causes Tangier disease, a rare HDL-deficiency syndrome. Whether heterozygosity for genetic variation in ABCA1 also contributes to HDL cholesterol (HDL-C) levels in the general population is presently unclear. We determined whether mutations...

  15. Cholesterol activates the G-protein coupled receptor Smoothened to promote Hedgehog signaling

    Science.gov (United States)

    Luchetti, Giovanni; Sircar, Ria; Kong, Jennifer H; Nachtergaele, Sigrid; Sagner, Andreas; Byrne, Eamon FX; Covey, Douglas F; Siebold, Christian; Rohatgi, Rajat

    2016-01-01

    Cholesterol is necessary for the function of many G-protein coupled receptors (GPCRs). We find that cholesterol is not just necessary but also sufficient to activate signaling by the Hedgehog (Hh) pathway, a prominent cell-cell communication system in development. Cholesterol influences Hh signaling by directly activating Smoothened (SMO), an orphan GPCR that transmits the Hh signal across the membrane in all animals. Unlike many GPCRs, which are regulated by cholesterol through their heptahelical transmembrane domains, SMO is activated by cholesterol through its extracellular cysteine-rich domain (CRD). Residues shown to mediate cholesterol binding to the CRD in a recent structural analysis also dictate SMO activation, both in response to cholesterol and to native Hh ligands. Our results show that cholesterol can initiate signaling from the cell surface by engaging the extracellular domain of a GPCR and suggest that SMO activity may be regulated by local changes in cholesterol abundance or accessibility. DOI: http://dx.doi.org/10.7554/eLife.20304.001 PMID:27705744

  16. Super-resolution microscopy reveals functional organization of dopamine transporters into cholesterol and neuronal activity-dependent nanodomains

    DEFF Research Database (Denmark)

    Rahbek-Clemmensen, Troels; Lycas, Matthew D.; Erlendsson, Simon

    2017-01-01

    to cholesterol depletion. Live photoactivated localization microscopy shows a similar dopamine transporter membrane organization in live heterologous cells. In neurons, dual-color dSTORM shows that tyrosine hydroxylase and vesicular monoamine transporter-2 are distinctively localized adjacent to...

  17. Absence of intestinal microbiota increases ß-cyclodextrin stimulated reverse cholesterol transport.

    Science.gov (United States)

    Mistry, Rima H; Verkade, Henkjan J; Tietge, Uwe J F

    2017-05-01

    Non-digestible oligosaccharides are used as prebiotics for perceived health benefits, among these modulating lipid metabolism. However, the mechanisms of action are incompletely understood. The present study characterized the impact of dietary ß-cyclodextrin (ßCD, 10%, w/w), a cyclic oligosaccharide, on sterol metabolism and reverse cholesterol transport (RCT) in conventional and also germ-free mice to establish dependency on metabolism by intestinal bacteria. In conventional ßCD-fed C57BL/6J wild-type mice plasma cholesterol decreased significantly (-40%, p < 0.05), largely within HDL, while fecal neutral sterol excretion increased (3-fold, p < 0.01) and fecal bile acid excretion was unchanged. Hepatic cholesterol levels and biliary cholesterol secretion were unaltered. Changes in cholesterol metabolism translated into increased macrophage-to-feces RCT in ßCD-administered mice (1.5-fold, p < 0.05). In germ-free C57BL/6J mice ßCD similarly lowered plasma cholesterol (-40%, p < 0.05). However, ßCD increased fecal neutral sterol excretion (7.5-fold, p < 0.01), bile acid excretion (2-fold, p < 0.05) and RCT (2.5-fold, p < 0.01) even more substantially in germ-free mice compared with the effect in conventional mice. In summary, this study demonstrates that ßCD lowers plasma cholesterol levels and increases fecal cholesterol excretion from a RCT-relevant pool. Intestinal bacteria decrease the impact of ßCD on RCT. These data suggest that dietary ßCD might have cardiovascular health benefits. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Intracellular transport of low density lipoprotein-derived cholesterol is defective in Niemann-Pick type C fibroblasts

    International Nuclear Information System (INIS)

    Liscum, L.; Ruggiero, R.M.; Faust, J.R.

    1989-01-01

    Niemann-Pick disease type C (NPC) is characterized by substantial intracellular accumulation of unesterified cholesterol. The accumulation of unesterified cholesterol in NPC fibroblasts cultured with low density lipoprotein (LDL) appears to result from the inability of LDL to stimulate cholesterol esterification in addition to impaired LDL-mediated downregulation of LDL receptor activity and cellular cholesterol synthesis. Although a defect in cholesterol transport in NPC cells has been inferred from previous studies, no experiments have been reported that measure the intracellular movement of LDL-cholesterol specifically. We have used four approaches to assess intracellular cholesterol transport in normal and NPC cells and have determined the following: (a) mevinolin-inhibited NPC cells are defective in using LDL-cholesterol for growth. However, exogenously added mevalonate restores cell growth equally in normal and NPC cells; (b) the transport of LDL-derived [3H]cholesterol to the plasma membrane is slower in NPC cells, while the rate of appearance of [3H]acetate-derived, endogenously synthesized [3H]cholesterol at the plasma membrane is the same for normal and NPC cells; (c) in NPC cells, LDL-derived [3H]cholesterol accumulates in lysosomes to higher levels than normal, resulting in defective movement to other cell membranes; and (d) incubation of cells with LDL causes an increase in cholesterol content of NPC lysosomes that is threefold greater than that observed in normal lysosomes. Our results indicate that a cholesterol transport defect exists in NPC that is specific for LDL-derived cholesterol

  19. Dietary egg white protein inhibits lymphatic lipid transport in thoracic lymph duct-cannulated rats.

    Science.gov (United States)

    Matsuoka, Ryosuke; Shirouchi, Bungo; Kawamura, Sayaka; Baba, Sanae; Shiratake, Sawako; Nagata, Kazuko; Imaizumi, Katsumi; Sato, Masao

    2014-11-05

    Dietary egg white protein (EWP) decreases serum cholesterol levels. We previously showed that EWP decreased cholesterol absorption in the intestine. Rats subjected to permanent lymph duct cannulation were used to investigate the effects of dietary EWP on lipid transport. They were fed diets with 20% EWP and casein, and their lymph was collected to quantify lymphatic lipid levels. Dietary EWP decreased lymphatic cholesterol transport compared with casein. It was previously shown that EWP excluded cholesterol from bile acid micelles. Therefore, pepsin-hydrolyzed EWP and casein were prepared. EWP was not completely digested. Ovalbumin, which is the most abundant protein in EWP, showed resistance to digestion by pepsin. This study investigated the effects of EWP pepsin hydrolysate (EWP-ph) on cholesterol micellar solubility, cholesterol transfer from the micellar to the oil phase, water-holding capacity (WHC), settling volume in water (SV), and relative viscosity and compared them with the effects of casein pepsin hydrolysate (C-ph). EWP-ph significantly decreased the micellar solubility and transfer rate and increased the WHC, SV, and relative viscosity compared with C-ph. Moreover, the pepsin hydrolysate of ovalbumin, a major protein in EWP, played a role in decreasing cholesterol micellar solubility, leading to the inhibition of cholesterol absorption. In conclusion, dietary EWP decreased cholesterol intestinal absorption by exerting combined effects of these physicochemical properties in the gut.

  20. Cholesterol transport and steroidogenesis by the corpus luteum

    Directory of Open Access Journals (Sweden)

    Christenson Lane K

    2003-11-01

    Full Text Available Abstract The synthesis of progesterone by the corpus luteum is essential for the establishment and maintenance of early pregnancy. Regulation of luteal steroidogenesis can be broken down into three major events; luteinization (i.e., conversion of an ovulatory follicle, luteal regression, and pregnancy induced luteal maintenance/rescue. While the factors that control these events and dictate the final steroid end products are widely varied among different species, the composition of the corpus luteum (luteinized thecal and granulosa cells and the enzymes and proteins involved in the steroidogenic pathway are relatively similar among all species. The key factors involved in luteal steroidogenesis and several new exciting observations regarding regulation of luteal steroidogenic function are discussed in this review.

  1. Tritium labelling of a cholesterol amphiphile designed for cell membrane anchoring of proteins.

    Science.gov (United States)

    Schäfer, Balázs; Orbán, Erika; Kele, Zoltán; Tömböly, Csaba

    2015-01-01

    Cell membrane association of proteins can be achieved by the addition of lipid moieties to the polypeptide chain, and such lipid-modified proteins have important biological functions. A class of cell surface proteins contains a complex glycosylphosphatidylinositol (GPI) glycolipid at the C-terminus, and they are accumulated in cholesterol-rich membrane microdomains, that is, lipid rafts. Semisynthetic lipoproteins prepared from recombinant proteins and designed lipids are valuable probes and model systems of the membrane-associated proteins. Because GPI-anchored proteins can be reinserted into the cell membrane with the retention of the biological function, they are appropriate candidates for preparing models via reduction of the structural complexity. A synthetic headgroup was added to the 3β-hydroxyl group of cholesterol, an essential lipid component of rafts, and the resulting cholesterol derivative was used as a simplified GPI mimetic. In order to quantitate the membrane integrated GPI mimetic after the exogenous addition to live cells, a tritium labelled cholesterol anchor was prepared. The radioactive label was introduced into the headgroup, and the radiolabelled GPI mimetic anchor was obtained with a specific activity of 1.37 TBq/mmol. The headgroup labelled cholesterol derivative was applied to demonstrate the sensitive detection of the cell membrane association of the anchor under in vivo conditions. Copyright © 2015 John Wiley & Sons, Ltd.

  2. Effects of Dietary Flavonoids on Reverse Cholesterol Transport, HDL Metabolism, and HDL Function.

    Science.gov (United States)

    Millar, Courtney L; Duclos, Quinn; Blesso, Christopher N

    2017-03-01

    Strong experimental evidence confirms that HDL directly alleviates atherosclerosis. HDL particles display diverse atheroprotective functions in reverse cholesterol transport (RCT), antioxidant, anti-inflammatory, and antiapoptotic processes. In certain inflammatory disease states, however, HDL particles may become dysfunctional and proatherogenic. Flavonoids show the potential to improve HDL function through their well-documented effects on cellular antioxidant status and inflammation. The aim of this review is to summarize the basic science and clinical research examining the effects of dietary flavonoids on RCT and HDL function. Based on preclinical studies that used cell culture and rodent models, it appears that many flavonoids (e.g., anthocyanidins, flavonols, and flavone subclasses) influence RCT and HDL function beyond simple HDL cholesterol concentration by regulating cellular cholesterol efflux from macrophages and hepatic paraoxonase 1 expression and activity. In clinical studies, dietary anthocyanin intake is associated with beneficial changes in serum biomarkers related to HDL function in a variety of human populations (e.g., in those who are hyperlipidemic, hypertensive, or diabetic), including increased HDL cholesterol concentration, as well as HDL antioxidant and cholesterol efflux capacities. However, clinical research on HDL functionality is lacking for some flavonoid subclasses (e.g., flavanols, flavones, flavanones, and isoflavones). Although there has been a tremendous effort to develop HDL-targeted drug therapies, more research is warranted on how the intake of foods or specific nutrients affects HDL function. © 2017 American Society for Nutrition.

  3. Model of OSBP-Mediated Cholesterol Supply to Aichi Virus RNA Replication Sites Involving Protein-Protein Interactions among Viral Proteins, ACBD3, OSBP, VAP-A/B, and SAC1.

    Science.gov (United States)

    Ishikawa-Sasaki, Kumiko; Nagashima, Shigeo; Taniguchi, Koki; Sasaki, Jun

    2018-04-15

    Positive-strand RNA viruses, including picornaviruses, utilize cellular machinery for genome replication. Previously, we reported that each of the 2B, 2BC, 2C, 3A, and 3AB proteins of Aichi virus (AiV), a picornavirus, forms a complex with the Golgi apparatus protein ACBD3 and phosphatidylinositol 4-kinase IIIβ (PI4KB) at viral RNA replication sites (replication organelles [ROs]), enhancing PI4KB-dependent phosphatidylinositol 4-phosphate (PI4P) production. Here, we demonstrate AiV hijacking of the cellular cholesterol transport system involving oxysterol-binding protein (OSBP), a PI4P-binding cholesterol transfer protein. AiV RNA replication was inhibited by silencing cellular proteins known to be components of this pathway, OSBP, the ER membrane proteins VAPA and VAPB (VAP-A/B), the PI4P-phosphatase SAC1, and PI-transfer protein β. OSBP, VAP-A/B, and SAC1 were present at RNA replication sites. We also found various previously unknown interactions among the AiV proteins (2B, 2BC, 2C, 3A, and 3AB), ACBD3, OSBP, VAP-A/B, and SAC1, and the interactions were suggested to be involved in recruiting the component proteins to AiV ROs. Importantly, the OSBP-2B interaction enabled PI4P-independent recruitment of OSBP to AiV ROs, indicating preferential recruitment of OSBP among PI4P-binding proteins. Protein-protein interaction-based OSBP recruitment has not been reported for other picornaviruses. Cholesterol was accumulated at AiV ROs, and inhibition of OSBP-mediated cholesterol transfer impaired cholesterol accumulation and AiV RNA replication. Electron microscopy showed that AiV-induced vesicle-like structures were close to ER membranes. Altogether, we conclude that AiV directly recruits the cholesterol transport machinery through protein-protein interactions, resulting in formation of membrane contact sites between the ER and AiV ROs and cholesterol supply to the ROs. IMPORTANCE Positive-strand RNA viruses utilize host pathways to modulate the lipid composition of

  4. PSRC1 overexpression attenuates atherosclerosis progression in apoE-/-mice by modulating cholesterol transportation and inflammation.

    Science.gov (United States)

    Guo, Kai; Hu, Lu; Xi, Dan; Zhao, Jinzhen; Liu, Jichen; Luo, Tiantian; Ma, Yusheng; Lai, Wenyan; Guo, Zhigang

    2018-03-01

    Human genome-wide association studies (GWAS) have found that proline/serine-rich coiled-coil 1 (PSRC1) encodes a protein that is associated with serum lipid levels and coronary artery disease. In addition, our previous study showed that the cholesterol efflux capacity is decreased in macrophages following a treatment silencing Psrc1, indicating that PSRC1 has anti-atherosclerotic effects. However, the role of PSRC1 in the development of atherosclerosis is unknown. This study aims to explore the effect of PSRC1 on atherosclerosis and its underlying mechanisms. A recombinant adenovirus expressing Psrc1 (Ad-PSRC1) was constructed and transfected in RAW264.7 cells as well as injected intravenously into apoE -/- mice. The in vitro study showed that PSRC1 overexpression reduced the cellular cholesterol content, increased the cholesterol efflux capacity and inhibited foam cell formation by upregulating the expression of peroxisome proliferator-activated receptor γ (PPAR-γ) and liver X receptor α (LXR-α), which are key cholesterol transportation-related proteins. Infecting apoE -/- mice with Ad-PSRC1 inhibited the development of atherosclerotic lesions and enhanced atherosclerotic plaque stability. Consistent with these results, PSRC1 overexpression in apoE -/- mice decreased the plasma levels of TC, TG, LDL-C, TNF-α, IL-1β and IL-6, increased the plasma HDL-C levels and improved HDL function. Similarly, the PPAR-γ and LXR-α expression levels were upregulated in the liver and in peritoneal macrophages of PSRC1-overexpressing apoE -/- mice. Finally, the liver and peritoneal macrophages of apoE -/- mice displayed elevated expression of β-catenin, which is a direct downstream gene of PSRC1 and an upstream gene of PPAR-γ and LXR-α, but decreased activity of nuclear transcription factor (NF-κB), which acts as a key gene in the regulation of inflammation. PSRC1 protects against the development of atherosclerosis and enhances the stability of plaques by modulating

  5. Following Intracellular Cholesterol Transport by Linear and Non-Linear Optical Microscopy of Intrinsically Fluorescent Sterols

    DEFF Research Database (Denmark)

    Wustner, D.

    2012-01-01

    Elucidation of intracellular cholesterol transport is important for understanding the molecular basis of several metabolic and neuronal diseases, like atheroclerosis or lysosomal storage disorders. Progress in this field depends crucially on the development of new technical approaches to follow...... the cellular movement of this essential lipid molecule. In this article, a survey of the various methods being used for analysis of sterol trafficking is given. Various classical biochemical methods are presented and their suitability for analysis of sterol trafficking is assessed. Special emphasis...... analysis like pixel-wise bleach rate fitting and multiphoton image correlation spectroscopy are introduced. Several applications of the new technology including observation of vectorial sterol trafficking in polarized human hepatoma cells for investigation of reverse cholesterol transport are presented....

  6. Cholesterol transport by the placenta : Placental liver X receptor activity as a modulator of fetal cholesterol metabolism?

    NARCIS (Netherlands)

    Plosch, T.; van Straten, E. M. E.; Kuipers, F.

    Cholesterol is an important sterol in mammals. Defects in cholesterol synthesis or intracellular routing have devastating consequences already in utero: the Smith-Lemli-Opitz syndrome, desmosterolosis and Niemann-Pick C I disease provide examples of severe human inherited diseases caused by

  7. Protein and cholesterol electrophoresis of plasma samples from captive cownose ray (Rhinoptera bonasus).

    Science.gov (United States)

    Cray, Carolyn; Rodriguez, Marilyn; Field, Cara; McDermott, Alexa; Leppert, Lynda; Clauss, Tonya; Bossart, Gregory D

    2015-11-01

    Our study was undertaken to assess the application of semiautomated methods available at the reference laboratory level for the evaluation of plasma protein and cholesterol via electrophoresis in samples from cownose rays (Rhinoptera bonasus). Three groups of animals were assessed: clinically normal, clinically abnormal, and parasitized with leeches. As reported previously, the albumin band was negligible; the protein electrophoretograms were dominated by a large beta-globulin fraction. While the group of samples from the leech-parasitized rays did not show any large differences, the abnormal group exhibited significantly elevated total solids and cholesterol levels. The latter was related to a significant increase in very low density lipoprotein levels. The results demonstrate the potential application of these laboratory methods in quantitation of plasma proteins and cholesterol fractions in subclass Elasmobranchii. © 2015 The Author(s).

  8. Inhibition of soluble epoxide hydrolase in mice promotes reverse cholesterol transport and regression of atherosclerosis.

    Science.gov (United States)

    Shen, Li; Peng, Hongchun; Peng, Ran; Fan, Qingsong; Zhao, Shuiping; Xu, Danyan; Morisseau, Christophe; Chiamvimonvat, Nipavan; Hammock, Bruce D

    2015-04-01

    Adipose tissue is the body largest free cholesterol reservoir and abundantly expresses ATP binding cassette transporter A1 (ABCA1), which maintains plasma high-density lipoprotein (HDL) levels. HDLs have a protective role in atherosclerosis by mediating reverse cholesterol transport (RCT). Soluble epoxide hydrolase (sEH) is a cytosolic enzyme whose inhibition has various beneficial effects on cardiovascular disease. The sEH is highly expressed in adipocytes, and it converts epoxyeicosatrienoic acids (EETs) into less bioactive dihydroxyeicosatrienoic acids. We previously showed that increasing EETs levels with a sEH inhibitor (sEHI) (t-AUCB) resulted in elevated ABCA1 expression and promoted ABCA1-mediated cholesterol efflux from 3T3-L1 adipocytes. The present study investigates the impacts of t-AUCB in mice deficient for the low density lipoprotein (LDL) receptor (Ldlr(-/-) mice) with established atherosclerotic plaques. The sEH inhibitor delivered in vivo for 4 weeks decreased the activity of sEH in adipose tissue, enhanced ABCA1 expression and cholesterol efflux from adipose depots, and consequently increased HDL levels. Furthermore, t-AUCB enhanced RCT to the plasma, liver, bile and feces. It also showed the reduction of plasma LDL-C levels. Consistently, t-AUCB-treated mice showed reductions in the size of atherosclerotic plaques. These studies establish that raising adipose ABCA1 expression, cholesterol efflux, and plasma HDL levels with t-AUCB treatment promotes RCT, decreasing LDL-C and atherosclerosis regression, suggesting that sEH inhibition may be a promising strategy to treat atherosclerotic vascular disease. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  9. Common Variants in Cholesterol Synthesis- and Transport-Related Genes Associate with Circulating Cholesterol Responses to Intakes of Conventional Dairy Products in Healthy Individuals.

    Science.gov (United States)

    Abdullah, Mohammad Mh; Cyr, Audrey; Lépine, Marie-Claude; Eck, Peter K; Couture, Patrick; Lamarche, Benoît; Jones, Peter Jh

    2016-05-01

    Dairy intake has been associated with varying impacts on circulating cholesterol concentrations across nutritional epidemiology and intervention studies, with findings attributed mainly to differences in the nature of dairy products consumed or study designs. The contribution of the genomic architecture to such observations has yet to be revealed. We assessed the impact of multiple common genetic variations in cholesterol-related genes on responses of serum cholesterol to the recommended amount of dairy product intake in Canada. In a multicenter, randomized crossover design, 101 normolipidemic adults (n = 29 men and 72 women), with a mean ± SD age of 41.7 ± 16.7 y and a body mass index (BMI, in kg/m(2)) of 25.9 ± 4.3 consumed 3 servings/d of dairy [375 mL 1% milk-fat (MF) milk, 175 g 1.5% MF yogurt, and 30 g of 34% MF cheese] or energy-matched control products (juice, cashews, and cookies) provided within a prudent background diet for 4 wk each, separated by a 4- to 8-wk washout period. Serum lipid variables were determined by standard enzymatic methods by using an autoanalyzer. Candidate single nucleotide polymorphisms were assessed by TaqMan genotyping assay. The responsiveness of serum total cholesterol (TC) and LDL cholesterol to the dairy compared with the control diet was associated with individuals' genotypes. The cholesterol transport gene ATP-binding cassette subfamily G, member 5 (ABCG5) rs6720173-GG homozygotes had higher concentrations of TC (+0.18 mmol/L; P = 0.0118) and LDL cholesterol (+0.17 mmol/L; P = 0.0056) relative to C-allele carriers (-0.07 and -0.06 mmol/L, respectively). The bile acid synthesis gene cholesterol 7α-hydroxylase (CYP7A1) rs3808607-G-allele carriers had higher TC (+0.20 to +0.28 mmol/L; P = 0.0026) and LDL cholesterol (+0.19 mmol/L for GT genotype; P = 0.0260) relative to TT homozygotes (-0.11 and -0.03 mmol/L, respectively). In addition, the cholesterol synthesis gene 7-dehydrocholesterol reductase (DHCR7) rs760241-A

  10. Cholesterol-induced protein sorting: an analysis of energetic feasibility

    DEFF Research Database (Denmark)

    Lundbaek, J A; Andersen, O S; Werge, T

    2003-01-01

    The mechanism(s) underlying the sorting of integral membrane proteins between the Golgi complex and the plasma membrane remain uncertain because no specific Golgi retention signal has been found. Moreover one can alter a protein's eventual localization simply by altering the length of its transme...

  11. Total protein and cholesterol concentrations in brain regions of male ...

    African Journals Online (AJOL)

    The results showed similarities (P>0.05) between the treatments in total protein concentrations in the cerebral cortex, medulla, hypothalamus, amygdala, mesencephalon and hippocampus. Total protein concentrations however differed significantly between diets (P<0.05) in the cerebellum and pons varoli with the lowest ...

  12. Cholesterol Transporters ABCA1 and ABCG1 Gene Expression in Peripheral Blood Mononuclear Cells in Patients with Metabolic Syndrome

    Directory of Open Access Journals (Sweden)

    Zahra Tavoosi

    2015-01-01

    Full Text Available ABCA1 and ABCG1 genes encode the cholesterol transporter proteins that play a key role in cholesterol and phospholipids homeostasis. This study was aimed at evaluating and comparing ABCA1 and ABCG1 genes expression in metabolic syndrome patients and healthy individuals. This case-control study was performed on 36 patients with metabolic syndrome and the same number of healthy individuals in Hamadan (west of Iran during 2013-2014. Total RNA was extracted from mononuclear cells and purified using RNeasy Mini Kit column. The expression of ABCA1 and ABCG1 genes was performed by qRT-PCR. Lipid profile and fasting blood glucose were measured using colorimetric procedures. ABCG1 expression in metabolic syndrome patients was significantly lower (about 75% compared to that of control group, while for ABCA1 expression, there was no significant difference between the two studied groups. Comparison of other parameters such as HDL-C, FBS, BMI, waist circumference, and systolic and diastolic blood pressure between metabolic syndrome patients and healthy individuals showed significant differences (P<0.05. Decrease in ABCG1 expression in metabolic syndrome patients compared to healthy individuals suggests that hyperglycemia, related metabolites, and hyperlipidemia over the transporter capacity resulted in decreased expression of ABCG1. Absence of a significant change in ABCA1 gene expression between two groups can indicate a different regulation mechanism for ABCA1 expression.

  13. Hepatic cholesterol transporter ABCG8 polymorphisms in gallstone disease in an Indian population.

    Science.gov (United States)

    Siddapuram, Siva Prasad; Mahurkar, Swapna; Duvvuru, Nageshwar Reddy; Mitnala, Sasikala; Guduru, Venkat Rao; Rebala, Pradeep; Mansard, Magnus Jayaraj

    2010-06-01

    Gallstone formation is characterized by the abnormal regulation of cholesterol trafficking and solubilization. The prevalence of gallstone disease (GSD) differs between ethnic groups sharing the common environment. These differences can be explained by a genetic predisposition to gallstone formation. Studies have identified single nucleotide polymorphisms (SNP) D19H and T400K in the cholesterol transporter gene ATP-binding cassette, subfamily G, member 8 (ABCG8) in patients with cholesterol gallstones. The aim of this study was to analyze the relationship between D19H and T400K polymorphisms in the ABCG8 gene and GSD in an Indian population, and the effects of these polymorphisms on cholesterol levels in sera and bile. A total of 226 patients with GSD were analyzed for their lipid profile in plasma and bile. A total of 289 controls were recruited, and their plasma lipid profile was analyzed by standard protocols. The genotype of SNP D19H and T400K of ABCG8 was analyzed in 226 patients and 222 control samples. SNP D19H was analyzed by direct sequencing, and SNP T400K genotyping was assayed by the amplification refractory mutation system-polymerase chain reaction. There was no significant difference in the allelic distribution of SNP T400K between the GSD and gallstone-free groups (P > 0.05), but the distribution of the SNP variant, D19H, was significantly higher (P = 0.017, odds ratio = 2.274) in patients compared to controls. The analysis of serum and bile cholesterol followed a strong association with genotypes. SNP D19H, but not SNP T400K, in the ABCG8 gene is significantly associated with GSD in an Indian population.

  14. The liver-selective thyromimetic T-0681 influences reverse cholesterol transport and atherosclerosis development in mice.

    Directory of Open Access Journals (Sweden)

    Ivan Tancevski

    2010-01-01

    Full Text Available Liver-selective thyromimetics have been reported to efficiently reduce plasma cholesterol through the hepatic induction of both, the low-density lipoprotein receptor (LDLr and the high-density lipoprotein (HDL receptor; the scavenger receptor class B type I (SR-BI. Here, we investigated the effect of the thyromimetic T-0681 on reverse cholesterol transport (RCT and atherosclerosis, and studied the underlying mechanisms using different mouse models, including mice lacking LDLr, SR-BI, and apoE, as well as CETP transgenic mice.T-0681 treatment promoted bile acid production and biliary sterol secretion consistently in the majority of the studied mouse models, which was associated with a marked reduction of plasma cholesterol. Using an assay of macrophage RCT in mice, we found T-0681 to significantly increase fecal excretion of macrophage-derived neutral and acidic sterols. No positive effect on RCT was found in CETP transgenic mice, most likely due to the observed decrease in plasma CETP mass. Studies in SR-BI KO and LDLr KO mice suggested hepatic LDLr to be necessary for the action of T-0681 on lipid metabolism, as the compound did not have any influence on plasma cholesterol levels in mice lacking this receptor. Finally, prolonged treatment with T-0681 reduced the development of atherosclerosis by 60% in apoE KOs on Western type diet. In contrast, at an earlier time-point T-0681 slightly increased small fatty streak lesions, in part due to an impaired macrophage cholesterol efflux capacity, when compared to controls.The present results show that liver-selective thyromimetics can promote RCT and that such compounds may protect from atherosclerosis partly through induction of bile acid metabolism and biliary sterol secretion. On-going clinical trials will show whether selective thyromimetics do prevent atherosclerosis also in humans.

  15. Transgenic overexpression of Niemann-Pick C2 protein promotes cholesterol gallstone formation in mice

    NARCIS (Netherlands)

    Acuna, Mariana; Gonzalez-Hodar, Lila; Amigo, Ludwig; Castro, Juan; Gabriela Morales, M.; Cancino, Gonzalo I.; Groen, Albert K.; Young, Juan; Francisco Miquel, Juan; Zanlungo, Silvana

    Background & Aims: Niemann-Pick C2 (NPC2) is a lysosomal protein involved in the egress of low-density lipoprotein-derived cholesterol from lysosomes to other intracellular compartments. NPC2 has been detected in several tissues and is also secreted from the liver into bile. We have previously shown

  16. Insig proteins mediate feedback inhibition of cholesterol synthesis in the intestine.

    Science.gov (United States)

    McFarlane, Matthew R; Liang, Guosheng; Engelking, Luke J

    2014-01-24

    Enterocytes are the only cell type that must balance the de novo synthesis and absorption of cholesterol, although the coordinate regulation of these processes is not well understood. Our previous studies demonstrated that enterocytes respond to the pharmacological blockade of cholesterol absorption by ramping up de novo sterol synthesis through activation of sterol regulatory element-binding protein-2 (SREBP-2). Here, we genetically disrupt both Insig1 and Insig2 in the intestine, two closely related proteins that are required for the feedback inhibition of SREBP and HMG-CoA reductase (HMGR). This double knock-out was achieved by generating mice with an intestine-specific deletion of Insig1 using Villin-Cre in combination with a germ line deletion of Insig2. Deficiency of both Insigs in enterocytes resulted in constitutive activation of SREBP and HMGR, leading to an 11-fold increase in sterol synthesis in the small intestine and producing lipidosis of the intestinal crypts. The intestine-derived cholesterol accumulated in plasma and liver, leading to secondary feedback inhibition of hepatic SREBP2 activity. Pharmacological blockade of cholesterol absorption was unable to further induce the already elevated activities of SREBP-2 or HMGR in Insig-deficient enterocytes. These studies confirm the essential role of Insig proteins in the sterol homeostasis of enterocytes.

  17. Following Intracellular Cholesterol Transport by Linear and Non-Linear Optical Microscopy of Intrinsically Fluorescent Sterols

    DEFF Research Database (Denmark)

    Wustner, D.

    2012-01-01

    analysis like pixel-wise bleach rate fitting and multiphoton image correlation spectroscopy are introduced. Several applications of the new technology including observation of vectorial sterol trafficking in polarized human hepatoma cells for investigation of reverse cholesterol transport are presented....... the cellular movement of this essential lipid molecule. In this article, a survey of the various methods being used for analysis of sterol trafficking is given. Various classical biochemical methods are presented and their suitability for analysis of sterol trafficking is assessed. Special emphasis...

  18. 13-hydroxy linoleic acid increases expression of the cholesterol transporters ABCA1, ABCG1 and SR-BI and stimulates apoA-I-dependent cholesterol efflux in RAW264.7 macrophages

    Directory of Open Access Journals (Sweden)

    Kämmerer Ines

    2011-11-01

    Full Text Available Abstract Background Synthetic activators of peroxisome proliferator-activated receptors (PPARs stimulate cholesterol removal from macrophages through PPAR-dependent up-regulation of liver × receptor α (LXRα and subsequent induction of cholesterol exporters such as ATP-binding cassette transporter A1 (ABCA1 and scavenger receptor class B type 1 (SR-BI. The present study aimed to test the hypothesis that the hydroxylated derivative of linoleic acid (LA, 13-HODE, which is a natural PPAR agonist, has similar effects in RAW264.7 macrophages. Methods RAW264.7 macrophages were treated without (control or with LA or 13-HODE in the presence and absence of PPARα or PPARγ antagonists and determined protein levels of LXRα, ABCA1, ABCG1, SR-BI, PPARα and PPARγ and apolipoprotein A-I mediated lipid efflux. Results Treatment of RAW264.7 cells with 13-HODE increased PPAR-transactivation activity and protein concentrations of LXRα, ABCA1, ABCG1 and SR-BI when compared to control treatment (P Conclusion 13-HODE induces cholesterol efflux from macrophages via the PPAR-LXRα-ABCA1/SR-BI-pathway.

  19. Drug transport proteins in the liver

    NARCIS (Netherlands)

    Faber, Klaas Nico; Muller, M.; Jansen, P.LM

    2003-01-01

    Together with drug metabolising enzymes, transmembrane transporters are important determinants of drug metabolism and drug clearance by the liver. Hepatic uptake of organic anions, cations, prostaglandins and bile salts is supported by dedicated transporter proteins in the basolateral (sinusoidal)

  20. ROLE OF ELEVATED LECITHIN - CHOLESTEROL ACYLTRANSFERASE AND CHOLESTERYL ESTER TRANSFER PROTEIN ACTIVITIES IN ABNORMAL LIPOPROTEINS FROM PROTEINURIC PATIENTS

    NARCIS (Netherlands)

    DULLAART, RPF; GANSEVOORT, RT; DIKKESCHEI, BD; DEZEEUW, D; DEJONG, PE; VANTOL, A

    Lecithin-cholesterol acyltransferase (LCAT) and cholesteryl ester transfer protein (CETP) are key factors in the esterification of free cholesterol, and the distribution of cholesteryl ester among lipoproteins in plasma. Alterations in these processes may play a role in the lipoprotein abnormalities

  1. Calcium-induced contraction and contractile protein of gallbladder smooth muscle after high-cholesterol feeding of prairie dogs

    NARCIS (Netherlands)

    Li, Y. F.; Weisbrodt, N. W.; Moody, F. G.; Coelho, J. C.; Gouma, D. J.

    1987-01-01

    Feeding a high-cholesterol diet to prairie dogs causes a reduction in contractile responses of gallbladder smooth muscle from these animals. In this study, the influence of cholesterol feeding on the contractile response to calcium and on the concentration of the contractile proteins actin and

  2. In situ mouse carotid perfusion model: glucose and cholesterol transport in the eye and brain.

    Science.gov (United States)

    Cattelotte, Julie; André, Pascal; Ouellet, Mélissa; Bourasset, Fanchon; Scherrmann, Jean-Michel; Cisternino, Salvatore

    2008-08-01

    The in situ mouse brain perfusion method for measuring blood-brain barrier permeability was adapted to assess transport of solutes at the blood-brain and blood-eye barriers. The procedure was checked with radiolabeled markers in oxygenated bicarbonate-buffered fluid infused for 30 to 120 sec via a carotid artery. Vascular flow estimated with diazepam was 2.2-fold lower in the eye than in the brain. The vascular volume and the integrity markers sucrose and inulin indicated that a perfusion flow rate of 2.5 mL/min preserved the physical integrity of these organs. However, the brain vasculature integrity was more sensitive to acute perfusion pressure than the eye vasculature. The functional capacities of blood barriers were assessed with D-glucose; its transport followed Michaelis-Menten kinetics with an apparent K(m) of 7.6 mmol/L and a V(max) of 23 micromol/sec per g in the brain, and a K(m) of 22.9 mmol/L and a V(max) of 40 micromol/sec per g in the eye. The transport of cholesterol to the brain and eye was significantly enhanced by adding the Abca1 inhibitor probucol, suggesting an Abca1-mediated efflux at the mouse brain and eye blood barriers. Thus in situ carotid perfusion is suitable for elucidating transport processes at the blood-brain and blood-eye barriers.

  3. Genetic variation in ABC transporter A1 contributes to HDL cholesterol in the general population

    DEFF Research Database (Denmark)

    Frikke-Schmidt, Ruth; Nordestgaard, Børge G; Jensen, Gorm B

    2004-01-01

    or single-nucleotide polymorphisms (SNPs) in ABCA1 were overrepresented in individuals with the lowest 1% (n=95) or highest 1% (n=95) HDL-C levels in the general population by screening the core promoter and coding region of ABCA1. For all nonsynonymous SNPs identified, we determined the effect of genotype......Homozygosity for mutations in ABC transporter A1 (ABCA1) causes Tangier disease, a rare HDL-deficiency syndrome. Whether heterozygosity for genetic variation in ABCA1 also contributes to HDL cholesterol (HDL-C) levels in the general population is presently unclear. We determined whether mutations...... on lipid traits in 9,259 individuals from the general population. Heterozygosity for ABCA1 mutations was identified in 10% of individuals with low HDL-C only. Three of 6 nonsynonymous SNPs (V771M, V825I, and R1587K) were associated with increases or decreases in HDL-C in women in the general population...

  4. Mevinolin-induced changes in cholesterol synthesis and protein glycosylation in lymphocytes of hypercholesterolemics

    International Nuclear Information System (INIS)

    Goel, V.; Premkumar, N.D.; Ramachandran, C.K.; Melnykovych, G.; Dujovne, C.A.

    1987-01-01

    Mevinolin (lovastatin, MVN), a potent competitive inhibitor of HMG CoA reductase (HMGR), has proven to be an effective hypolipidemic agent in patients with non-homozygous primary hypercholesterolemia. Since inhibition of HMGR can also reduce the synthesis of non-sterol mevalonate products such as dolichols, it was of interest to examine the dolichol-mediated cellular reactions in MVN-treated patients. Blood was collected from patients after various durations of MVN therapy. Peripheral lymphocytes were isolated using Ficoll-Paque gradient. The cells were suspended in RPMI-1640 medium and pulsed in the presence of 14 C-2-acetate or 3 H-mannose for 30 min. At the end of incubation the radioactivity recovered in non-saponifiable fraction ( 14 C) or TCA precipitable protein ( 3 H) was measured. Cholesterol synthesis continued to fall gradually and remained low throughout, in direct correlation with falls in plasma LDL cholesterol levels. Incorporation of mannose into protein fraction was reduced by the 1st month of therapy, remained low until the 7th month and recovered by the 10th month while on MVN. In summary, MVN appears to reduce cholesterol synthesis continuously but its inhibitory effect on glycosylation seems to be overcome after prolonged therapy. This escape effect could result from a rebound increase in HMGR in response to its competitive inhibition by MVN

  5. IMB2026791, a Xanthone, Stimulates Cholesterol Efflux by Increasing the Binding of Apolipoprotein A-I to ATP-Binding Cassette Transporter A1

    Directory of Open Access Journals (Sweden)

    Zijian Xie

    2012-03-01

    Full Text Available It is known that the ATP-binding cassette transporter A1 (ABCA1 plays a major role in cholesterol homeostasis and high density lipoprotein (HDL metabolism. Several laboratories have demonstrated that ABCA1 binding to lipid-poor apolipoprotein A-I (apoA-I will mediate the assembly of nascent HDL and cellular cholesterol efflux, which suggests a possible receptor-ligand interaction between ABCA1 and apoA-I. In this study, a cell-based-ELISA-like high-throughput screening (HTS method was developed to identify the synthetic and natural compounds that can regulate binding activity of ABCA1 to apoA-I. The cell-based-ELISA-like high-throughput screen was conducted in a 96-well format using Chinese hamster ovary (CHO cells stably transfected with ABCA1 pIRE2-EGFP (Enhanced Green Fluorecence Protein expression vector and the known ABCA1 inhibitor glibenclamide as the antagonist control. From 2,600 compounds, a xanthone compound (IMB 2026791 was selected using this HTS assay, and it was proved as an apoA-I binding agonist to ABCA1 by a flow cytometry assay and western blot analysis. The [3H] cholesterol efflux assay of IMB2026791 treated ABCA1-CHO cells and PMA induced THP-1 macrophages (human acute monocytic leukemia cell further confirmed the compound as an accelerator of cholesterol efflux in a dose-dependent manner with an EC50 of 25.23 μM.

  6. The Role of Maternal-Fetal Cholesterol Transport in Early Fetal Life : Current Insights

    NARCIS (Netherlands)

    Baardman, Maria E.; Kerstjens-Frederikse, Wilhelmina S.; Berger, Rolf M. F.; Bakker, Marian K.; Hofstra, Robert M. W.; Plosch, Torsten

    The importance of maternal cholesterol as an exogenous cholesterol source for the growing embryo was first reported in studies of Smith-Lemli-Opitz syndrome. Although most of the fetus's cholesterol is synthesized by the fetus itself, there is now growing evidence that during the first weeks of

  7. The role of maternal-fetal cholesterol transport in early fetal life: Current insights

    NARCIS (Netherlands)

    T. Baardman (Taco); W.S. Kerstjens-Frederikse (Wilhelmina); R.M.F. Berger (Rolf); M.K. Bakker (Marian); R.M.W. Hofstra (Robert); T. Plösch (Torsten)

    2013-01-01

    textabstractThe importance of maternal cholesterol as an exogenous cholesterol source for the growing embryo was first reported in studies of Smith-Lemli-Opitz syndrome. Although most of the fetus's cholesterol is synthesized by the fetus itself, there is now growing evidence that during the first

  8. Synthesis and live-cell imaging of fluorescent sterols for analysis of intracellular cholesterol transport

    DEFF Research Database (Denmark)

    Modzel, Maciej; Lund, Frederik W.; Wüstner, Daniel

    2017-01-01

    Cellular cholesterol homeostasis relies on precise control of the sterol content of organelle membranes. Obtaining insight into cholesterol trafficking pathways and kinetics by live-cell imaging relies on two conditions. First, one needs to develop suitable analogs that resemble cholesterol...

  9. Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus.

    Science.gov (United States)

    Bukiya, Anna N; Durdagi, Serdar; Noskov, Sergei; Rosenhouse-Dantsker, Avia

    2017-04-14

    Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. N-terminal domain of the cholesterol transporter Niemann-Pick C1-like 1 (NPC1L1) is essential for α-tocopherol transport.

    Science.gov (United States)

    Kamishikiryo, Jun; Haraguchi, Misaki; Nakashima, Shunsuke; Tasaka, Yuka; Narahara, Hiroe; Sugihara, Narumi; Nakamura, Tetsuya; Morita, Tetsuo

    2017-04-29

    Both cholesterol and α-tocopherol are essential lipophilic nutrients for humans and animals. Although cholesterol in excess causes severe problems such as coronary heart disease, it is a necessary component of cell membranes and is the precursor for the biosynthesis of steroid hormones and bile acids. Niemann-Pick C1-like 1 (NPC1L1) is a cholesterol transporter that is highly expressed in the small intestine and liver in humans and plays an important role in cholesterol homeostasis. Cholesterol promotes NPC1L1 endocytosis, which is an early step in cholesterol uptake. Furthermore, α-tocopherol is the most active form of vitamin E, and sufficient amounts of vitamin E are critical for health. It has been reported that NPC1L1 mediates α-tocopherol absorption; however, the mechanisms underlying this process are unknown. In this study, we found that treatment of cells that stably express NPC1L1-GFP with α-tocopherol promotes NPC1L1 endocytosis, and the NPC1L1 inhibitor, ezetimibe, efficiently prevents the α-tocopherol-induced endocytosis of NPC1L1. Cholesterol binding to the N-terminal domain (NTD) of NPC1L1 (NPC1L1-NTD) is essential for NPC1L1-mediated cholesterol absorption. We found that α-tocopherol competitively binds NPC1L1-NTD with cholesterol. Furthermore, when cells stably expressed NPC1L1ΔNTD-GFP, α-tocopherol could not induce the endocytosis of NPC1L1ΔNTD. Taken together, these results demonstrate that NPC1L1 recognizes α-tocopherol via its NTD and mediates α-tocopherol uptake through the same mechanism as cholesterol absorption. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. The association of 83 Plasma proteins with CHD mortality, BMI, HDL-, and total cholesterol in men: applying multivariate statistics to identify proteins with prognostic value and biological relevance

    NARCIS (Netherlands)

    Heidema, A.G.; Thissen, U.; Boer, J.M.; Bouwman, F.G.; Feskens, E.J.M.; Mariman, E.C.

    2009-01-01

    In this study, we applied the multivariate statistical tool Partial Least Squares (PLS) to analyze the relative importance of 83 plasma proteins in relation to coronary heart disease (CHD) mortality and the intermediate end points body mass index, HDL-cholesterol and total cholesterol. From a Dutch

  12. A mixture design approach to optimizing low cholesterol mayonnaise formulation prepared with wheat germ protein isolate.

    Science.gov (United States)

    Rahbari, Mahshid; Aalami, Mehran; Kashaninejad, Mahdi; Maghsoudlou, Yahya; Aghdaei, Seid Soheil Amiri

    2015-06-01

    The aim of this study was to optimize the mixture proportion of low cholesterol mayonnaise containing wheat germ protein isolate (WGPI) and xanthan gum (XG), as emulsifying agents in mayonnaise preparation. The mayonnaise prepared with different combinations of WGPI, egg yolk (0-9 % of each component) and XG (0-0.5 %). The optimized mixture proportions of low cholesterol mayonnaise were determined by applying the optimal mixture design method to acquire the mayonnaise with proper stability, texture, rheological properties and sensory scores. Optimum values of WGPI, XG and egg yolk in the mixture were found to be 7.87 %, 0.2 % and 0.93 %, respectively (of 9 % egg yolk). The WGPI, due to unique functional properties, had the greatest effect on properties of mayonnaise samples. Moreover, combination of XG and WGPI, improved the stability, heat stability, viscosity, consistency coefficient and textural properties of product. However, the overall acceptance was maximum in a mixture contained high amount of WGPI and XG and low amount of egg yolk. The results of this research showed the feasibility of preparation a low cholesterol mayonnaise by application a desirable combination of WGPI, XG, and egg yolk, with comparable properties those of the conventional mayonnaise.

  13. Control of Insulin Granule Formation and Function by the ABC Transporters ABCG1 and ABCA1 and by Oxysterol Binding Protein OSBP.

    Science.gov (United States)

    Hussain, Syed Saad; Harris, Megan T; Kreutzberger, Alex J B; Inouye, Candice M; Doyle, Catherine A; Castle, Anna M; Arvan, Peter; Castle, J David

    2018-03-14

    In pancreatic β cells, insulin granule membranes are enriched in cholesterol and are both recycled and newly generated. Cholesterol's role in supporting granule membrane formation and function is poorly understood. A TP b inding c assette transporters ABCG1 and ABCA1 regulate intracellular cholesterol and are important for insulin secretion. RNAi-induced depletion in cultured pancreatic β cells shows that ABCG1 is needed to stabilize newly made insulin granules against lysosomal degradation; ABCA1 is also involved but to a lesser extent. Both transporters are also required for optimum glucose-stimulated insulin secretion, likely via complementary roles. Exogenous cholesterol addition rescues knockdown-induced granule loss (ABCG1) and reduced secretion (both transporters). Another cholesterol transport protein, oxysterol binding protein (OSBP) appears to act proximally as a source of endogenous cholesterol for granule formation. Its knockdown caused similar defective stability of young granules and glucose-stimulated insulin secretion, neither of which were rescued with exogenous cholesterol. Dual knockdowns of OSBP and ABC transporters support their serial function in supplying and concentrating cholesterol for granule formation. OSBP knockdown also decreased proinsulin synthesis consistent with a proximal ER defect. Thus, membrane cholesterol distribution contributes to insulin homeostasis at production, packaging and export levels through the actions of OSBP and ABCs G1 and A1. © 2018 by The American Society for Cell Biology.

  14. The cholesterol-lowering effects of oat varieties based on their difference in the composition of proteins and lipids.

    Science.gov (United States)

    Guo, Lina; Tong, Li-Tao; Liu, Liya; Zhong, Kui; Qiu, Ju; Zhou, Sumei

    2014-12-05

    The aim of present study is to investigate the hypocholesterolemic effects of the oat components other than the β-glucan in rats fed with a hypercholesterolemic diet. Four-week-old male Wister rats were divided into 6 groups of 7 rats each with similar mean body weights and serum cholesterol concentrations. Rats were fed with the experimental diets containing 10% oats flour for 30 days. Food intake was recorded and monitored everyday to ensure the similar contents of protein, starch, lipid and cellulose in all groups. The lipids levels in serum, liver, and faeces were determined. The plasma total cholesterol concentrations in different oat groups were significantly reduced compared with the control group, and the effects were different among oat groups. The decrease extent of plasma total cholesterol and low-density lipoprotein-cholesterol concentrations increased with the increase of the proteins and lipids contents. Moreover, liver total cholesterol and cholesterol ester contents were markedly decreased. The fecal bile acids concentrations in the oat groups were significantly increased. Oat proteins had lower Lysine/Arginin (0.59 ~ 0.66) and Methionin/Glycine (0.27 ~ 0.35) ratio than casein (Lysine/Arginin, 2.33; Methionin/Glycine, 1.51). Oat lipids contained higher contents of total Vitamin E and plant sterols than that in soybean oil. These results indicated that dietary oat improved hypercholesterolemia by increasing the excretions of fecal bile acids, and this improvement was not only related to β-glucan, but also attributed to the lipids and proteins. Oat proteins decreased serum total cholesterol and low-density lipoprotein-cholesterol contents due to their low Lysine/Arginin and Methionin/Glycine ratio. The co-existence of oleic acid, linoleic, vitamin E, or plant sterols accounted for the hypocholesterolemic properties of oat lipids.

  15. Na,K-ATPase reconstituted in ternary liposome: the presence of cholesterol affects protein activity and thermal stability.

    Science.gov (United States)

    Yoneda, Juliana Sakamoto; Rigos, Carolina Fortes; de Lourenço, Thaís Fernanda Aranda; Sebinelli, Heitor Gobbi; Ciancaglini, Pietro

    2014-12-15

    Differential scanning calorimetry (DSC) was applied to investigate the effect of cholesterol on the thermotropic properties of the lipid membrane (DPPC and DPPE). The thermostability and unfolding of solubilized and reconstituted Na,K-ATPase in DPPC:DPPE:cholesterol-liposomes was also studied to gain insight into the role of cholesterol in the Na,K-ATPase modulation of enzyme function and activity. The tertiary system (DPPC:DPPE:cholesterol) (molar ratio DPPC:DPPE equal 1:1) when cholesterol content was increased from 0% up to 40% results in a slight decrease in the temperature of transition and enthalpy, and an increase in width. We observed that, without heating treatment, at 37°C, the activity was higher for 20mol% cholesterol. However, thermal inactivation experiments showed that the enzyme activity loss time depends on the cholesterol membrane content. The unfolding of the enzyme incorporated to liposomes of DPPC:DPPE (1:1mol) with different cholesterol contents, ranging from 0% to 40% mol was also studied by DSC. Some differences between the thermograms indicate that the presence of lipids promotes a conformational change in protein structure and this change is enough to change the way Na,K-ATPase thermally unfolds. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Regulation of cellular transport by klotho protein.

    Science.gov (United States)

    Sopjani, Mentor; Rinnerthaler, Mark; Almilaji, Ahmad; Ahmeti, Salih; Dermaku-Sopjani, Miribane

    2014-01-01

    The antiaging protein of Klotho is a transmembrane protein mainly expressed in the kidney, parathyroid glands and choroid plexus of the brain. The Klotho protein exists in two forms, a full-length membrane form and a soluble secreted form. The extracellular domain of Klotho can be enzymatically cleaved off and released into the systemic circulation where it acts as β-glucuronidase and a hormone. Soluble Klotho can be found in the blood, cerebrospinal fluid, and the urine of mammals. Klotho deficiency results in early appearance of multiple age-related disorders and premature death, whereas overexpression of Klotho exerts the opposite effect. Klotho may influence cellular transport processes across the cell membrane by inhibiting calcitriol (1,25(OH) (2)D(3)), formation or by directly affecting transporter proteins, including ion channels, carriers and pumps. Accordingly, Klotho protein is a powerful regulator of transport mechanisms across the cell membrane. Klotho regulates diverse calcium and potassium ion channels, as well as several carriers including the Na(+)-coupled excitatory amino acid transporters EAAT3 and EAAT4, the Na(+)-coupled phosphate cotransporters, NaPi-IIa and NaPi-IIb, and a Na(+)/K(+)-ATPase. All those cellular transport regulations contribute in the aging suppressor role of Klotho. Future studies will help to determine if the Klotho protein regulates cell-surface expression of other transport proteins and is affecting underlying mechanisms.

  17. The Dietary Constituent Falcarindiol Promotes Cholesterol Efflux from THP-1 Macrophages by Increasing ABCA1 Gene Transcription and Protein Stability

    OpenAIRE

    Wang, Limei; Palme, Veronika; Schilcher, Nicole; Ladurner, Angela; Heiss, Elke H.; Stangl, Herbert; Bauer, Rudolf; Dirsch, Verena M.; Atanasov, Atanas G.

    2017-01-01

    We report increased cholesterol efflux from macrophages in the presence of falcarindiol, an important dietary constituent present in commonly used vegetables and medicinal plants. Falcarindiol (3–20 μM) increased cholesterol efflux from THP-1-derived macrophages. Western blot analysis showed an increased protein level of ABCA1 upon falcarindiol exposure. Quantitative real-time PCR revealed that also ABCA1 mRNA level rise with falcarindiol (10 μM) treatment. The effect of falcarindiol on ABCA1...

  18. The morphology and composition of cholesterol-rich micellar nanostructures determine transmembrane protein (GPCR) activity.

    Science.gov (United States)

    O'Malley, Michelle A; Helgeson, Matthew E; Wagner, Norman J; Robinson, Anne S

    2011-01-19

    We examined model mixed micelles consisting of the nonionic surfactant n-dodecyl-β-D-maltoside, 3-(3-cholamidopropyl)-dimethylammoniopropane sulfonate, and the cholesterol derivative cholesteryl hemisuccinate (CHS) to identify micellar properties that are correlated with the in vitro conformational stability and activity of the human adenosine A₂a receptor, a G-protein coupled receptor. Small-angle neutron scattering was used to determine micellar structure and composition as a function of concentration of the various components, and radioligand binding was used as a sensitive probe for receptor activity. Micelles adopted an oblate ellipsoidal morphology and exhibited a reduction in size and change in curvature upon addition of CHS. Our results show a strong correlation between the number of CHS monomers per micelle and the activity of the receptor reconstituted in those micelles. Micelles that yield optimal human adenosine A₂a receptor stability closely mimic the cholesterol composition and thickness of mammalian membranes. Thus, successful reconstitution of the receptor is dependent on both specific lipid-protein interactions and the geometry of the micelle environment. Copyright © 2011 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  19. Inhibition of protein kinase C affects on mode of synaptic vesicle exocytosis due to cholesterol depletion

    Energy Technology Data Exchange (ETDEWEB)

    Petrov, Alexey M., E-mail: fysio@rambler.ru; Zakyrjanova, Guzalija F., E-mail: guzik121192@mail.ru; Yakovleva, Anastasia A., E-mail: nastya1234qwer@mail.ru; Zefirov, Andrei L., E-mail: zefiroval@rambler.ru

    2015-01-02

    Highlights: • We examine the involvement of PKC in MCD induced synaptic vesicle exocytosis. • PKC inhibitor does not decrease the effect MCD on MEPP frequency. • PKC inhibitor prevents MCD induced FM1-43 unloading. • PKC activation may switch MCD induced exocytosis from kiss-and-run to a full mode. • Inhibition of phospholipase C does not lead to similar change in exocytosis. - Abstract: Previous studies demonstrated that depletion of membrane cholesterol by 10 mM methyl-beta-cyclodextrin (MCD) results in increased spontaneous exocytosis at both peripheral and central synapses. Here, we investigated the role of protein kinase C in the enhancement of spontaneous exocytosis at frog motor nerve terminals after cholesterol depletion using electrophysiological and optical methods. Inhibition of the protein kinase C by myristoylated peptide and chelerythrine chloride prevented MCD-induced increases in FM1-43 unloading, whereas the frequency of spontaneous postsynaptic events remained enhanced. The increase in FM1-43 unloading still could be observed if sulforhodamine 101 (the water soluble FM1-43 quencher that can pass through the fusion pore) was added to the extracellular solution. This suggests a possibility that exocytosis of synaptic vesicles under these conditions could occur through the kiss-and-run mechanism with the formation of a transient fusion pore. Inhibition of phospholipase C did not lead to similar change in MCD-induced exocytosis.

  20. Regulation of the membrane insertion and conductance activity of the metamorphic chloride intracellular channel protein CLIC1 by cholesterol.

    Directory of Open Access Journals (Sweden)

    Stella M Valenzuela

    Full Text Available The Chloride Intracellular ion channel protein CLIC1 has the ability to spontaneously insert into lipid membranes from a soluble, globular state. The precise mechanism of how this occurs and what regulates this insertion is still largely unknown, although factors such as pH and redox environment are known contributors. In the current study, we demonstrate that the presence and concentration of cholesterol in the membrane regulates the spontaneous insertion of CLIC1 into the membrane as well as its ion channel activity. The study employed pressure versus area change measurements of Langmuir lipid monolayer films; and impedance spectroscopy measurements using tethered bilayer membranes to monitor membrane conductance during and following the addition of CLIC1 protein. The observed cholesterol dependent behaviour of CLIC1 is highly reminiscent of the cholesterol-dependent-cytolysin family of bacterial pore-forming proteins, suggesting common regulatory mechanisms for spontaneous protein insertion into the membrane bilayer.

  1. Cholesterol Corrects Altered Conformation of MHC-II Protein in Leishmania donovani Infected Macrophages: Implication in Therapy

    Science.gov (United States)

    Chakrabarti, Saikat; Roy, Syamal

    2016-01-01

    Background Previously we reported that Kala-azar patients show progressive decrease in serum cholesterol as a function of splenic parasite burden. Splenic macrophages (MΦ) of Leishmania donovani (LD) infected mice show decrease in membrane cholesterol, while LD infected macrophages (I-MΦ) show defective T cell stimulating ability that could be corrected by liposomal delivery of cholesterol. T helper cells recognize peptide antigen in the context of class II MHC molecule. It is known that the conformation of a large number of membrane proteins is dependent on membrane cholesterol. In this investigation we tried to understand the influence of decreased membrane cholesterol in I-MΦ on the conformation of MHC-II protein and peptide-MHC-II stability, and its bearing on the antigen specific T-cell activation. Methodology/Principal Findings MΦ of CBA/j mice were infected with Leishmania donovani (I-MΦ). Two different anti-Aκ mAbs were used to monitor the status of MHC-II protein under parasitized condition. One of them (11.5–2) was conformation specific, whereas the other one (10.2.16) was not. Under parasitized condition, the binding of 11.5–2 decreased significantly with respect to the normal counterpart, whereas that of 10.2.16 remained unaltered. The binding of 11.5–2 was restored to normal upon liposomal delivery of cholesterol in I-MΦ. By molecular dynamics (MD) simulation studies we found that there was considerable conformational fluctuation in the transmembrane domain of the MHC-II protein in the presence of membrane cholesterol than in its absence, which possibly influenced the distal peptide binding groove. This was evident from the faster dissociation of the cognate peptide from peptide-MHC complex under parasitized condition, which could be corrected by liposomal delivery of cholesterol in I-MΦ. Conclusion The decrease in membrane cholesterol in I-MΦ may lead to altered conformation of MHC II, and this may contribute to a faster dissociation of

  2. HDL cholesterol response to GH replacement is associated with common cholesteryl ester transfer protein gene variation (-629C>A) and modified by glucocorticoid treatment

    NARCIS (Netherlands)

    Dullaart, Robin P. F.; van den Berg, Gerrit; van der Knaap, Aafke M.; Dijck-Brouwer, Janneke; Dallinga-Thie, Geesje M.; Zelissen, Peter M. J.; Sluiter, Wim J.; van Beek, André P.

    2010-01-01

    GH replacement lowers total cholesterol and low-density lipoprotein cholesterol (LDL-C) in GH-deficient adults, but effects on high-density lipoprotein (HDL) cholesterol (HDL-C) are variable. Both GH and glucocorticoids decrease cholesteryl ester transfer protein (CETP) activity, which is important

  3. Modulation of microRNA Expression in Subjects with Metabolic Syndrome and Decrease of Cholesterol Efflux from Macrophages via microRNA-33-Mediated Attenuation of ATP-Binding Cassette Transporter A1 Expression by Statins.

    Science.gov (United States)

    Chen, Wei-Ming; Sheu, Wayne H-H; Tseng, Pei-Chi; Lee, Tzong-Shyuan; Lee, Wen-Jane; Chang, Pey-Jium; Chiang, An-Na

    2016-01-01

    Metabolic syndrome (MetS) is a complicated health problem that encompasses a variety of metabolic disorders. In this study, we analyzed the relationship between the major biochemical parameters associated with MetS and circulating levels of microRNA (miR)-33, miR-103, and miR-155. We found that miRNA-33 levels were positively correlated with levels of fasting blood glucose, glycosylated hemoglobin A1c, total cholesterol, LDL-cholesterol, and triacylglycerol, but negatively correlated with HDL-cholesterol levels. In the cellular study, miR-33 levels were increased in macrophages treated with high glucose and cholesterol-lowering drugs atorvastatin and pitavastatin. miR-33 has been reported to play an essential role in cholesterol homeostasis through ATP-binding cassette transporter A1 (ABCA1) regulation and reverse cholesterol transport. However, the molecular mechanism underlying the linkage between miR-33 and statin treatment remains unclear. In the present study, we investigated whether atorvastatin and pitavastatin exert their functions through the modulation of miR-33 and ABCA1-mediated cholesterol efflux from macrophages. The results showed that treatment of the statins up-regulated miR-33 expression, but down-regulated ABCA1 mRNA levels in RAW264.7 cells and bone marrow-derived macrophages. Statin-mediated ABCA1 regulation occurs at the post-transcriptional level through targeting of the 3'-UTR of the ABCA1 transcript by miR-33. Additionally, we found significant down-regulation of ABCA1 protein expression in macrophages treated with statins. Finally, we showed that high glucose and statin treatment significantly suppressed cholesterol efflux from macrophages. These findings have highlighted the complexity of statins, which may exert detrimental effects on metabolic abnormalities through regulation of miR-33 target genes.

  4. Modulation of microRNA Expression in Subjects with Metabolic Syndrome and Decrease of Cholesterol Efflux from Macrophages via microRNA-33-Mediated Attenuation of ATP-Binding Cassette Transporter A1 Expression by Statins.

    Directory of Open Access Journals (Sweden)

    Wei-Ming Chen

    Full Text Available Metabolic syndrome (MetS is a complicated health problem that encompasses a variety of metabolic disorders. In this study, we analyzed the relationship between the major biochemical parameters associated with MetS and circulating levels of microRNA (miR-33, miR-103, and miR-155. We found that miRNA-33 levels were positively correlated with levels of fasting blood glucose, glycosylated hemoglobin A1c, total cholesterol, LDL-cholesterol, and triacylglycerol, but negatively correlated with HDL-cholesterol levels. In the cellular study, miR-33 levels were increased in macrophages treated with high glucose and cholesterol-lowering drugs atorvastatin and pitavastatin. miR-33 has been reported to play an essential role in cholesterol homeostasis through ATP-binding cassette transporter A1 (ABCA1 regulation and reverse cholesterol transport. However, the molecular mechanism underlying the linkage between miR-33 and statin treatment remains unclear. In the present study, we investigated whether atorvastatin and pitavastatin exert their functions through the modulation of miR-33 and ABCA1-mediated cholesterol efflux from macrophages. The results showed that treatment of the statins up-regulated miR-33 expression, but down-regulated ABCA1 mRNA levels in RAW264.7 cells and bone marrow-derived macrophages. Statin-mediated ABCA1 regulation occurs at the post-transcriptional level through targeting of the 3'-UTR of the ABCA1 transcript by miR-33. Additionally, we found significant down-regulation of ABCA1 protein expression in macrophages treated with statins. Finally, we showed that high glucose and statin treatment significantly suppressed cholesterol efflux from macrophages. These findings have highlighted the complexity of statins, which may exert detrimental effects on metabolic abnormalities through regulation of miR-33 target genes.

  5. Comparison of serum leptin, glucose, total cholesterol and total protein levels in fertile and repeat breeder cows

    Directory of Open Access Journals (Sweden)

    Saime Guzel

    2014-12-01

    Full Text Available In the present study we measured serum glucose, leptin, total cholesterol and total protein concentrations in repeat breeder cows and compared them with fertile cows. For this aim, 20 repeat breeder cows and 20 fertile cows were used as material. Repeat breeder cows were found to have lower levels of leptin and glucose as compared with fertile ones. No significant differences in total cholesterol and total protein levels were observed between the two groups. No significant correlation of leptin with glucose, total cholesterol and total protein was observed in fertile and repeat breeder cows. Low concentrations of glucose and leptin can have some effects on reproductive problems as repeat breeder and help to understand potential mechanisms impairing fertility in repeat breeder cows.

  6. The cholesterol transporter ABCA1 is expressed in stallion spermatozoa and reproductive tract tissues.

    Science.gov (United States)

    Merkl, M; Ertl, R; Handschuh, S; Aurich, C; Schäfer-Somi, S

    2016-04-01

    strong signals in Leydig cells were present in prepubertal stallions. In prepubertal stallions, the ABCA1 messenger RNA level in testicular tissue was significantly higher than in adult stallions. We conclude that the ABCA1 transport molecule is present in adult and prepubertal stallion spermatozoa as well as testicular and epididymal tissue. ABCA1 is supposed to contribute to cholesterol transport and to support capacitation; however, this remains to be proven by functional studies. Species-specific differences concerning the localization inside the spermatozoa membrane are alike. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Phytosterols Differentially Influence ABC transporter Expression, Cholesterol Efflux and Inflammatory Cytokine Secretion in Macrophage Foam Cells

    Science.gov (United States)

    Sabeva, Nadezhda S; McPhaul, Christopher M; Li, Xiangan; Cory, Theodore J.; Feola, David J.; Graf, Gregory A

    2010-01-01

    Phytosterol supplements lower low density lipoprotein (LDL) cholesterol, but accumulate in vascular lesions of patients and limit the anti-atherosclerotic effects of LDL lowering in apolipoprotein E deficient mice, suggesting that the cholesterol lowering benefit of phytosterol supplementation may not be fully realized. Individual phytosterols have cell-type specific effects that may either be beneficial or deleterious with respect to atherosclerosis, but little is known concerning their effects on macrophage function. The effects of phytosterols on ABCA1 and ABCG1 abundance, cholesterol efflux, and inflammatory cytokine secretion were determined in cultured macrophage foam cells. Among the commonly consumed phytosterols, stigmasterol increased expression of ABCA1 and ABCG1 and increased efflux of cholesterol to apolipoprotein (Apo) AI and high density lipoprotein (HDL). Campesterol and sitosterol had no effect on ABCA1 or ABCG1 levels. Sitosterol had no effect of cholesterol efflux to Apo AI or HDL, whereas campesterol had a modest, but significant reduction in cholesterol efflux to HDL in THP-1 macrophages. Whereas stigmasterol blunted aggregated LDL-induced increases in tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1β secretion, sitosterol exacerbated these effects. The presence of campesterol had no effect on agLDL-induced inflammatory cytokine secretion from THP-1 macrophages. In conclusion, the presence of stigmasterol in modified lipoproteins promoted cholesterol efflux and suppressed inflammatory cytokine secretion in response to lipid loading in macrophage foam cells. While campesterol was largely inert, the presence of sitosterol increased the proinflammatory cytokine secretion. PMID:21111593

  8. Interleukin-10 increases reverse cholesterol transport in macrophages through its bidirectional interaction with liver X receptor α

    Energy Technology Data Exchange (ETDEWEB)

    Halvorsen, Bente, E-mail: Bente.Halvorsen@rr-research.no [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Institute of Clinical Medicine, University of Oslo, Oslo (Norway); K.G. Jebsen Inflammation Research Center, University of Oslo, Oslo (Norway); Holm, Sverre [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Yndestad, Arne [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Institute of Clinical Medicine, University of Oslo, Oslo (Norway); K.G. Jebsen Inflammation Research Center, University of Oslo, Oslo (Norway); Scholz, Hanne [Section for Transplantation, Institute for Surgical Research, Oslo University Hospital Rikshospitalet, Oslo (Norway); Sagen, Ellen Lund [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Nebb, Hilde [Department of Nutrition, Institute for Basic Medical Sciences, University of Oslo, Oslo (Norway); Institute of Clinical Medicine, University of Oslo, Oslo (Norway); Holven, Kirsten B. [Department of Nutrition, Institute for Basic Medical Sciences, University of Oslo, Oslo (Norway); Dahl, Tuva B. [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Institute of Clinical Medicine, University of Oslo, Oslo (Norway); Aukrust, Pål [Research Institute of Internal Medicine, Oslo University Hospital Rikshospitalet, Oslo (Norway); Section of Clinical Immunology and Infectious Diseases, Oslo University Hospital Rikshospitalet, Oslo (Norway); Institute of Clinical Medicine, University of Oslo, Oslo (Norway); K.G. Jebsen Inflammation Research Center, University of Oslo, Oslo (Norway)

    2014-08-08

    Highlights: • IL-10 promotes reverse cholesterol efflux from lipid loaded macrophages. • IL-10 increases the expression of ABCA-1 and ABCG-1. • IL-10 exhibits cross-talk with the nuclear receptor LXRα. - Abstract: Interleukin (IL)-10 is a prototypical anti-inflammatory cytokine that has been shown to attenuate atherosclerosis development. In addition to its anti-inflammatory properties, the anti-atherogenic effect of IL-10 has recently also been suggested to reflect a complex effect of IL-10 on lipid metabolism in macrophages. In the present study we examined the effects of IL-10 on cholesterol efflux mechanism in lipid-loaded THP-1 macrophages. Our main findings were: (i) IL-10 significantly enhanced cholesterol efflux induced by fetal-calf serum, high-density lipoprotein (HDL){sub 2} and apolipoprotein A-1. (ii) The IL-10-mediated effects on cholesterol efflux were accompanied by an increased IL-10-mediated expression of the ATP-binding cassette transporters ABCA1 and ABCG1, that was further enhanced when the cells were co-activated with the liver X receptor (LXR)α agonist (22R)-hydroxycholesterol. (iii) The effect of LXRα activation on the IL-10-mediated effects on the ATP-binding cassette transporters seems to include enhancing effects on the IL-10 receptor 1 (IL10R1) expression and interaction with STAT-3 signaling. (iv) These enhancing effects on ABCA1 and ABCG1 was not seen when the cells were stimulated with the IL-10 family members IL-22 and IL-24. This study suggests that the anti-atherogenic properties of IL-10 may include enhancing effects on cholesterol efflux mechanism that involves cross-talk with LXRα activation.

  9. Interleukin-10 increases reverse cholesterol transport in macrophages through its bidirectional interaction with liver X receptor α

    International Nuclear Information System (INIS)

    Halvorsen, Bente; Holm, Sverre; Yndestad, Arne; Scholz, Hanne; Sagen, Ellen Lund; Nebb, Hilde; Holven, Kirsten B.; Dahl, Tuva B.; Aukrust, Pål

    2014-01-01

    Highlights: • IL-10 promotes reverse cholesterol efflux from lipid loaded macrophages. • IL-10 increases the expression of ABCA-1 and ABCG-1. • IL-10 exhibits cross-talk with the nuclear receptor LXRα. - Abstract: Interleukin (IL)-10 is a prototypical anti-inflammatory cytokine that has been shown to attenuate atherosclerosis development. In addition to its anti-inflammatory properties, the anti-atherogenic effect of IL-10 has recently also been suggested to reflect a complex effect of IL-10 on lipid metabolism in macrophages. In the present study we examined the effects of IL-10 on cholesterol efflux mechanism in lipid-loaded THP-1 macrophages. Our main findings were: (i) IL-10 significantly enhanced cholesterol efflux induced by fetal-calf serum, high-density lipoprotein (HDL) 2 and apolipoprotein A-1. (ii) The IL-10-mediated effects on cholesterol efflux were accompanied by an increased IL-10-mediated expression of the ATP-binding cassette transporters ABCA1 and ABCG1, that was further enhanced when the cells were co-activated with the liver X receptor (LXR)α agonist (22R)-hydroxycholesterol. (iii) The effect of LXRα activation on the IL-10-mediated effects on the ATP-binding cassette transporters seems to include enhancing effects on the IL-10 receptor 1 (IL10R1) expression and interaction with STAT-3 signaling. (iv) These enhancing effects on ABCA1 and ABCG1 was not seen when the cells were stimulated with the IL-10 family members IL-22 and IL-24. This study suggests that the anti-atherogenic properties of IL-10 may include enhancing effects on cholesterol efflux mechanism that involves cross-talk with LXRα activation

  10. Transport proteins promoting Escherichia coli pathogenesis

    Science.gov (United States)

    Tang, Fengyi; Saier, Milton H.

    2014-01-01

    Escherichia coli is a genetically diverse species infecting hundreds of millions of people worldwide annually. We examined seven well-characterized E. coli pathogens causing urinary tract infections, gastroenteritis, pyelonephritis and haemorrhagic colitis. Their transport proteins were identified and compared with each other and a non-pathogenic E. coli K12 strain to identify transport proteins related to pathogenesis. Each pathogen possesses a unique set of protein secretion systems for export to the cell surface or for injecting effector proteins into host cells. Pathogens have increased numbers of iron siderophore receptors and ABC iron uptake transporters, but the numbers and types of low-affinity secondary iron carriers were uniform in all strains. The presence of outer membrane iron complex receptors and high-affinity ABC iron uptake systems correlated, suggesting co-evolution. Each pathovar encodes a different set of pore-forming toxins and virulence-related outer membrane proteins lacking in K12. Intracellular pathogens proved to have a characteristically distinctive set of nutrient uptake porters, different from those of extracellular pathogens. The results presented in this report provide information about transport systems relevant to various types of E. coli pathogenesis that can be exploited in future basic and applied studies. PMID:24747185

  11. Transport proteins promoting Escherichia coli pathogenesis.

    Science.gov (United States)

    Tang, Fengyi; Saier, Milton H

    2014-01-01

    Escherichia coli is a genetically diverse species infecting hundreds of millions of people worldwide annually. We examined seven well-characterized E. coli pathogens causing urinary tract infections, gastroenteritis, pyelonephritis and haemorrhagic colitis. Their transport proteins were identified and compared with each other and a non-pathogenic E. coli K12 strain to identify transport proteins related to pathogenesis. Each pathogen possesses a unique set of protein secretion systems for export to the cell surface or for injecting effector proteins into host cells. Pathogens have increased numbers of iron siderophore receptors and ABC iron uptake transporters, but the numbers and types of low-affinity secondary iron carriers were uniform in all strains. The presence of outer membrane iron complex receptors and high-affinity ABC iron uptake systems correlated, suggesting co-evolution. Each pathovar encodes a different set of pore-forming toxins and virulence-related outer membrane proteins lacking in K12. Intracellular pathogens proved to have a characteristically distinctive set of nutrient uptake porters, different from those of extracellular pathogens. The results presented in this report provide information about transport systems relevant to various types of E. coli pathogenesis that can be exploited in future basic and applied studies. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Synergistic activation of G protein-gated inwardly rectifying potassium channels by cholesterol and PI(4,5)P2.

    Science.gov (United States)

    Bukiya, Anna N; Rosenhouse-Dantsker, Avia

    2017-07-01

    G-protein gated inwardly rectifying potassium (GIRK or Kir3) channels play a major role in the control of the heart rate, and require the membrane phospholipid phosphatidylinositol-bis-phosphate (PI(4,5)P 2 ) for activation. Recently, we have shown that the activity of the heterotetrameric Kir3.1/Kir3.4 channel that underlies atrial K ACh currents was enhanced by cholesterol. Similarly, the activities of both the Kir3.4 homomer and its active pore mutant Kir3.4* (Kir3.4_S143T) were also enhanced by cholesterol. Here we employ planar lipid bilayers to investigate the crosstalk between PI(4,5)P 2 and cholesterol, and demonstrate that these two lipids act synergistically to activate Kir3.4* currents. Further studies using the Xenopus oocytes heterologous expression system suggest that PI(4,5)P 2 and cholesterol act via distinct binding sites. Whereas PI(4,5)P 2 binds to the cytosolic domain of the channel, the putative binding region of cholesterol is located at the center of the transmembrane domain overlapping the central glycine hinge region of the channel. Together, our data suggest that changes in the levels of two key membrane lipids - cholesterol and PI(4,5)P 2 - could act in concert to provide fine-tuning of Kir3 channel function. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Computational comparison of a calcium-dependent jellyfish protein (apoaequorin) and calmodulin-cholesterol in short-term memory maintenance.

    Science.gov (United States)

    Morrill, Gene A; Kostellow, Adele B; Gupta, Raj K

    2017-03-06

    Memory reconsolidation and maintenance depend on calcium channels and on calcium/calmodulin-dependent kinases regulating protein turnover in the hippocampus. Ingestion of a jellyfish protein, apoaequorin, reportedly protects and/or improves verbal learning in adults and is currently widely advertised for use by the elderly. Apoaequorin is a member of the EF-hand calcium binding family of proteins that includes calmodulin. Calmodulin-1 (148 residues) differs from Apoaequorin (195 residues) in that it contains four rather than three Ca 2+ -binding sites and three rather than four cholesterol-binding (CRAC, CARC) domains. All three cholesterol-binding CARC domains in calmodulin have a high interaction affinity for cholesterol compared to only two high affinity CARC domains in apoaequorin. Both calmodulin and apoaequorin can form dimers with a potential of eight bound Ca 2+ ions and six high affinity-bound cholesterol molecules in calmodulin with six bound Ca 2+ ions and a mixed population of eight cholesterols bound to both CARC and CRAC domains in apoaqueorin. MEMSAT-SVM analysis indicates that both calmodulin and apoaqueorin have a pore-lining region. The Peptide-Cutter algorithm predicts that calmodulin-1 contains 11 trypsin-specific cleavage sites (compared to 21 in apoaqueorin), four of which are potentially blocked by cholesterol and three are within the Ca-binding domains and/or the pore-lining region. Three are clustered between the third and fourth Ca 2+ -binding sites. Only calmodulin pore-lining regions contain Ca 2+ binding sites and as dimers may insert into the plasma membrane of neural cells and act as Ca 2+ channels. In a dietary supplement, bound cholesterol may protect both apoaequorin and calmodulin from proteolysis in the gut as well as facilitate uptake across the blood-brain barrier. Our results suggest that a physiological calmodulin-cholesterol complex, not cholesterol-free jellyfish protein, may better serve as a dietary supplement to

  14. Soliton/exciton transport in proteins.

    Science.gov (United States)

    Sinkala, Zachariah

    2006-08-21

    The study of electron/proton transport in alpha-helix sections of proteins have illustrated the existence of soliton-like mechanisms. Recently, Ciblis and Cosic extended investigation to the existence of possible like soliton-type mechanisms in other parts of the protein. They used Quantum Hamiltonian analysis to investigate. In this paper, we investigate the same problem but we use Classical Hamiltonian analysis in our investigation.

  15. Cholesterol transporter ATP-binding cassette A1 (ABCA1) is elevated in prion disease and affects PrPC and PrPSc concentrations in cultured cells.

    Science.gov (United States)

    Kumar, Rajeev; McClain, Denise; Young, Rebecca; Carlson, George A

    2008-06-01

    Prion diseases are transmissible neurodegenerative disorders of prion protein (PrP) conformation. Prion replication by conversion of benign PrPC isoforms into disease-specific PrPSc isoforms is intimately involved in prion disease pathogenesis and may be initiated in cholesterol-rich caveolae-like domains (CLD). Concentrations of the cholesterol transporter ATP-binding cassette A1 protein (ABCA1) are elevated in pre-clinical scrapie prion-infected mice and in prion-infected cells in vitro. Elevation of ABCA1 in prion-infected brain is not a direct consequence of local PrPSc accumulation, indeed levels of ABCA1 are comparable in brain regions that differ dramatically in the amount of PrPSc. Similarly, ABCA1 concentrations are identical in normal mice, transgenic mice overexpressing PrP and PrP knockout mice. In contrast, PrPC and PrPSc levels, but not Prnp mRNA, were increased by overexpression of ABCA1 in N2a neuroblastoma cells and scrapie prion-infected N2a cells (ScN2a). Conversely, RNAi-mediated knock down of Abca1 expression decreased the concentrations of PrPC in N2a cells and of PrPSc in ScN2a cells. These results suggest that ABCA1's effects on PrPC levels are post-translational and may reflect an increase in of PrPC stability, mediated either indirectly by increasing membrane cholesterol and CLD formation or by other functions of ABCA1. The increased supply of PrPC available for conversion would lead to increased PrPSc formation.

  16. Complement Activation by Ceramide Transporter Proteins

    NARCIS (Netherlands)

    Bode, G.H.; Losen, M.; Buurman, W.A.; Veerhuis, R.; Molenaar, P.C.; Steinbusch, H.W.M.; De Baets, M.H.; Daha, MR; Martinez-Martinez, P.

    2014-01-01

    C1q is the initiator of the classical complement pathway and, as such, is essential for efficient opsonization and clearance of pathogens, altered self-structures, and apoptotic cells. The ceramide transporter protein (CERT) and its longer splicing isoform CERTL are known to interact with

  17. The Dietary Constituent Falcarindiol Promotes Cholesterol Efflux from THP-1 Macrophages by Increasing ABCA1 Gene Transcription and Protein Stability

    Directory of Open Access Journals (Sweden)

    Limei Wang

    2017-09-01

    Full Text Available We report increased cholesterol efflux from macrophages in the presence of falcarindiol, an important dietary constituent present in commonly used vegetables and medicinal plants. Falcarindiol (3–20 μM increased cholesterol efflux from THP-1-derived macrophages. Western blot analysis showed an increased protein level of ABCA1 upon falcarindiol exposure. Quantitative real-time PCR revealed that also ABCA1 mRNA level rise with falcarindiol (10 μM treatment. The effect of falcarindiol on ABCA1 protein as well as mRNA level were counteracted by co-treatment with BADGE, an antagonist of PPARγ. Furthermore, falcarindiol significantly inhibited ABCA1 protein degradation in the presence of cycloheximide. This post-translational regulation of ABCA1 by falcarindiol occurs most likely by inhibition of lysosomal cathepsins, resulting in decreased proteolysis and extended protein half-life of ABCA1. Taken together, falcarindiol increases ABCA1 protein level by two complementary mechanisms, i.e., promoting ABCA1 gene expression and inhibiting ABCA1 protein degradation, which lead to enhanced cholesterol efflux.

  18. Langmuir–Blodgett films of cholesterol oxidase and S-layer proteins onto screen-printed electrodes

    Energy Technology Data Exchange (ETDEWEB)

    Guimarães, Juliana Aguilar, E-mail: helen@peq.coppe.ufrj.br; Ferraz, Helen Conceição; Alves, Tito Lívio Moitinho

    2014-04-01

    Graphical abstract: - Highlights: • Langmuir and LB monolayers of ChOx and S-layer proteins were obtained. • Mixed ChOx/S-layer proteins films presented an ideal-like behavior. • Modified sensor showed stable peaks with moderate intensity. • The type of LB deposition affects the sensor ability of detecting cholesterol. • Mixed ChOx/S-layer proteins LB films improve sensor properties. - Abstract: Stable Langmuir monolayers of cholesterol oxidase (ChOx) and S-layer proteins were produced at the water–air interface and subsequently transferred onto the surface of screen-printed carbon electrodes by the Langmuir–Blodgett (LB) technique. The modified electrode surface was characterized by atomic force microscopy (AFM) and cyclic voltammetry (CV). AFM indicated the presence of deposited layers, showing reduction of surface roughness (RMS and Rt parameters). Significant changes in the shape of CVs were observed in modified electrodes compared to bare electrodes. The anodic peaks could be observed in cyclic voltammograms (CV), at a scan rate equal to 25 mV s{sup −1}, using electrodes with Z-type LB deposition. The presence of S-layer proteins in the ChOx LB film increases the oxidation peak intensity and reduces the oxidation potential. Altogether, these results demonstrate the feasibility of producing a cholesterol biosensor based on the immobilization of ChOx and S-layer proteins by LB technique.

  19. The Dietary Constituent Falcarindiol Promotes Cholesterol Efflux from THP-1 Macrophages by Increasing ABCA1 Gene Transcription and Protein Stability.

    Science.gov (United States)

    Wang, Limei; Palme, Veronika; Schilcher, Nicole; Ladurner, Angela; Heiss, Elke H; Stangl, Herbert; Bauer, Rudolf; Dirsch, Verena M; Atanasov, Atanas G

    2017-01-01

    We report increased cholesterol efflux from macrophages in the presence of falcarindiol, an important dietary constituent present in commonly used vegetables and medicinal plants. Falcarindiol (3-20 μM) increased cholesterol efflux from THP-1-derived macrophages. Western blot analysis showed an increased protein level of ABCA1 upon falcarindiol exposure. Quantitative real-time PCR revealed that also ABCA1 mRNA level rise with falcarindiol (10 μM) treatment. The effect of falcarindiol on ABCA1 protein as well as mRNA level were counteracted by co-treatment with BADGE, an antagonist of PPARγ. Furthermore, falcarindiol significantly inhibited ABCA1 protein degradation in the presence of cycloheximide. This post-translational regulation of ABCA1 by falcarindiol occurs most likely by inhibition of lysosomal cathepsins, resulting in decreased proteolysis and extended protein half-life of ABCA1. Taken together, falcarindiol increases ABCA1 protein level by two complementary mechanisms, i.e., promoting ABCA1 gene expression and inhibiting ABCA1 protein degradation, which lead to enhanced cholesterol efflux.

  20. Niemann-Pick Type C2 Protein Mediates Hepatic Stellate Cells Activation by Regulating Free Cholesterol Accumulation

    Directory of Open Access Journals (Sweden)

    Yuh-Ching Twu

    2016-07-01

    Full Text Available In chronic liver diseases, regardless of their etiology, the development of fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. Hepatic stellate cells (HSCs are the main profibrogenic cells that promote the pathogenesis of liver fibrosis, and so it is important to identify the molecules that regulate HSCs activation and liver fibrosis. Niemann-Pick type C2 (NPC2 protein plays an important role in the regulation of intracellular cholesterol homeostasis by directly binding with free cholesterol. However, the roles of NPC2 in HSCs activation and liver fibrosis have not been explored in detail. Since a high-cholesterol diet exacerbates liver fibrosis progression in both rodents and humans, we propose that the expression of NPC2 affects free cholesterol metabolism and regulates HSCs activation. In this study, we found that NPC2 is decreased in both thioacetamide- and carbon tetrachloride-induced liver fibrosis tissues. In addition, NPC2 is expressed in quiescent HSCs, but its activation status is down-regulated. Knockdown of NPC2 in HSC-T6 cells resulted in marked increases in transforming growth factor-β1 (TGF-β1-induced collagen type 1 α1 (Col1a1, α-smooth muscle actin (α-SMA expression, and Smad2 phosphorylation. In contrast, NPC2 overexpression decreased TGF-β1-induced HSCs activation. We further demonstrated that NPC2 deficiency significantly increased the accumulation of free cholesterol in HSCs, increasing Col1a1 and α-SMA expression and activating Smad2, and leading to sensitization of HSCs to TGF-β1 activation. In contrast, overexpression of NPC2 decreased U18666A-induced free cholesterol accumulation and inhibited the subsequent HSCs activation. In conclusion, our study has demonstrated that NPC2 plays an important role in HSCs activation by regulating the accumulation of free cholesterol. NPC2 overexpression may thus represent a new treatment strategy for liver fibrosis.

  1. Long-term ethanol consumption impairs reverse cholesterol transport function of high-density lipoproteins by depleting high-density lipoprotein sphingomyelin both in rats and in humans.

    Science.gov (United States)

    Marmillot, Philippe; Munoz, Jennifer; Patel, Sanket; Garige, Mamatha; Rosse, Richard B; Lakshman, M Raj

    2007-07-01

    Moderate alcohol consumption has been linked to lower incidence of coronary artery disease due to increased plasma high-density lipoprotein (HDL), whereas heavy drinking has the opposite effect. Because of the crucial role of HDL in reverse cholesterol transport and positive correlation of HDL sphingomyelin (SM) content with cholesterol efflux, we have compared HDL SM content with its reverse cholesterol transport capacity both in rats fed ethanol on long-term basis and alcoholic individuals. In rats, SM HDL content was decreased in the ethanol group (-15.4%, P cholesterol uptake with control-group hepatocytes and 35.0% (P cholesterol uptake with ethanol-group hepatocytes. Conversely, hepatocytes from the ethanol group, when compared with hepatocytes from the control group, exhibited 31.0% (P cholesterol uptake with control-group HDL and 48.0% (P alcoholic individuals without liver disease (-51.5%, P alcoholic individuals with liver disease (-51.3%, P alcoholic individuals without liver disease, both efflux and uptake were decreased by 83.0% and 54.0% (P alcoholic individuals with liver disease by 84.0% and 61.0% (P consumption significantly impairs not only cholesterol efflux function of HDL by decreasing its SM content but also cholesterol uptake by affecting presumably hepatocyte receptors for HDL.

  2. Genetic variation in the cholesterol transporter NPC1L1, ischaemic vascular disease, and gallstone disease

    DEFF Research Database (Denmark)

    Lauridsen, Bo Kobberø; Stender, Stefan; Frikke-Schmidt, Ruth

    2015-01-01

    developed IVD or symptomatic gallstone disease, respectively, during follow-up from 1977 to 2013. We genotyped four common NPC1L1 variants, previously associated with reduced LDL cholesterol levels, thus mimicking the effect of ezetimibe, and calculated a weighted genotype score. With increasing genotype...

  3. Cholesterol Promotes Protein Binding by Affecting Membrane Electrostatics and Solvation Properties.

    Science.gov (United States)

    Doktorova, Milka; Heberle, Frederick A; Kingston, Richard L; Khelashvili, George; Cuendet, Michel A; Wen, Yi; Katsaras, John; Feigenson, Gerald W; Vogt, Volker M; Dick, Robert A

    2017-11-07

    Binding of the retroviral structural protein Gag to the cellular plasma membrane is mediated by the protein's matrix (MA) domain. Prominent among MA-PM interactions is electrostatic attraction between the positively charged MA domain and the negatively charged plasma membrane inner leaflet. Previously, we reported that membrane association of HIV-1 Gag, as well as purified Rous sarcoma virus (RSV) MA and Gag, depends strongly on the presence of acidic lipids and is enhanced by cholesterol (Chol). The mechanism underlying this enhancement was unclear. Here, using a broad set of in vitro and in silico techniques we addressed molecular mechanisms of association between RSV MA and model membranes, and investigated how Chol enhances this association. In neutron scattering experiments with liposomes in the presence or absence of Chol, MA preferentially interacted with preexisting POPS-rich clusters formed by nonideal lipid mixing, binding peripherally to the lipid headgroups with minimal perturbation to the bilayer structure. Molecular dynamics simulations showed a stronger MA-bilayer interaction in the presence of Chol, and a large Chol-driven increase in lipid packing and membrane surface charge density. Although in vitro MA-liposome association is influenced by disparate variables, including ionic strength and concentrations of Chol and charged lipids, continuum electrostatic theory revealed an underlying dependence on membrane surface potential. Together, these results conclusively show that Chol affects RSV MA-membrane association by making the electrostatic potential at the membrane surface more negative, while decreasing the penalty for lipid headgroup desolvation. The presented approach can be applied to other viral and nonviral proteins. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  4. Cognition, learning behaviour and hippocampal synaptic plasticity are not disrupted in mice over-expressing the cholesterol transporter ABCG1

    Directory of Open Access Journals (Sweden)

    Eadie Brennan D

    2009-02-01

    Full Text Available Abstract Background Cognitive deficits are a hallmark feature of both Down Syndrome (DS and Alzheimer's Disease (AD. Extra copies of the genes on chromosome 21 may also play an important role in the accelerated onset of AD in DS individuals. Growing evidence suggests an important function for cholesterol in the pathogenesis of AD, particularly in APP metabolism and production of Aβ peptides. The ATP-Binding Cassette-G1 (ABCG1 transporter is located on chromosome 21, and participates in the maintenance of tissue cholesterol homeostasis. Results To assess the role of ABCG1 in DS-related cognition, we evaluated the cognitive performance of mice selectively over-expressing the ABCG1 gene from its endogenous regulatory signals. Both wild-type and ABCG1 transgenic mice performed equivalently on several behavioral tests, including measures of anxiety, as well as on reference and working memory tasks. No deficits in hippocampal CA1 synaptic plasticity as determined with electrophysiological studies were apparent in mice over-expressing ABCG1. Conclusion These findings indicate that although ABCG1 may play a role in maintaining cellular or tissue cholesterol homeostasis, it is unlikely that excess ABCG1 expression contributes to the cognitive deficits in DS individuals.

  5. Function of MRP1/ABCC1 is not dependent on cholesterol or cholesterol-stabilized lipid rafts

    NARCIS (Netherlands)

    Meszaros, Peter; Klappe, Karin; Hummel, Ina; Hoekstra, Dick; Kok, Jan Willem

    2011-01-01

    MRP1 (multidrug-resistance-related protein 1)/ABCC1 (ATP-binding cassette transporter C1) has been localized in cholesterol-enriched lipid rafts, which suggests a role for these lipid rafts and/or cholesterol in MRP1 function. In the present study, we have shown for the first time that nearly

  6. Effect of dietary omega-3 fatty acids and chronic ethanol consumption on reverse cholesterol transport in rats.

    Science.gov (United States)

    Marmillot, P; Rao, M N; Liu, Q H; Chirtel, S J; Lakshman, M R

    2000-04-01

    We previously showed that chronic ethanol feeding leads to a decrease of apolipoprotein E (apoE) in high-density lipoprotein (HDL), whereas supplementing this diet with 2.8% of total dietary calories as omega3-fatty acids (omega3FAs) restores HDL-apoE to the control values. Since HDL containing apoE plays a major role in reverse cholesterol transport (RCT), we measured the effects chronic ethanol intake and omega3-FAs on RCT in the present study. Four groups of rats, control normal fat (CN), alcohol-normal fat (AN), control omega3FA fat (CF), and alcohol-omega3FA fat (AF), were fed their respective diets for 8 weeks, after which hepatocytes and HDLs from each group were evaluated for RCT capacity (cholesterol efflux from macrophages and uptake by liver cells). Compared with the control diet (CN), chronic ethanol (AN) feeding inhibited the cholesterol efflux capacity of HDL by 21% (P cholesterol uptake by the liver, there were no significant 3-way or 4-way interactions between the 4 factors, HDL-alcohol, HDL-fish oil, hepatocyte-alcohol, and hepatocyte-fish oil. The main effects for HDL-alcohol, HDL-fish oil, and hepatocyte-alcohol were all highly significant (P = .0001, .0001, and .007, respectively). There was a significant HDL-alcohol and HDL-fish oil interaction (P = .0001). Hepatocyte-alcohol was not a factor in any 2-way interactions. Our study indicates no evidence of an interaction between the effects of omega3FAs and the effects of alcohol on hepatocytes in terms of RCT function. Thus, feeding as little as 2.8% of the total dietary calories as omega3FA not only restored the impaired RCT function of HDL caused by chronic ethanol intake, but also enhanced by severalfold the ability of HDL to promote RCT even in normal animals.

  7. Eicosapentaenoic acid membrane incorporation impairs ABCA1-dependent cholesterol efflux via a protein kinase A signaling pathway in primary human macrophages.

    Science.gov (United States)

    Fournier, Natalie; Tardivel, Sylviane; Benoist, Jean-François; Vedie, Benoît; Rousseau-Ralliard, Delphine; Nowak, Maxime; Allaoui, Fatima; Paul, Jean-Louis

    2016-04-01

    A diet rich in n-3/n-6 polyunsaturated fatty acids (PUFAs) is cardioprotective. Dietary PUFAs affect the cellular phospholipids composition, which may influence the function of membrane proteins. We investigated the impact of the membrane incorporation of several PUFAs on ABCA1-mediated cholesterol efflux, a key antiatherogenic pathway. Arachidonic acid (AA) (C20:4 n-6) and docosahexaenoic acid (DHA) (C22:6 n-3) decreased or increased cholesterol efflux from J774 mouse macrophages, respectively, whereas they had no effect on efflux from human monocyte-derived macrophages (HMDM). Importantly, eicosapentaenoic acid (EPA) (C20:5 n-3) induced a dose-dependent reduction of ABCA1 functionality in both cellular models (-28% for 70μM of EPA in HMDM), without any alterations in ABCA1 expression. These results show that PUFA membrane incorporation does not have the same consequences on cholesterol efflux from mouse and human macrophages. The EPA-treated HMDM exhibited strong phospholipid composition changes, with high levels of both EPA and its elongation product docosapentaenoic acid (DPA) (C22:5 n-3), which is associated with a decreased level of AA. In HMDM, EPA reduced the ATPase activity of the membrane transporter. Moreover, the activation of adenylate cyclase by forskolin and the inhibition of cAMP phosphodiesterase by isobutylmethylxanthine restored ABCA1 cholesterol efflux in EPA-treated human macrophages. In conclusion, EPA membrane incorporation reduces ABCA1 functionality in mouse macrophages as well as in primary human macrophages and this effect seems to be PKA-dependent in human macrophages. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. miR-92a enhances recombinant protein productivity in CHO cells by increasing intracellular cholesterol levels.

    Science.gov (United States)

    Loh, Wan Ping; Yang, Yuansheng; Lam, Kong Peng

    2017-04-01

    MicroRNAs (miRNAs) have emerged as promising targets for engineering of CHO cell factories to enhance recombinant protein productivity. Manipulation of miRNA levels in CHO cells have been shown to improve product yield by increasing proliferation and specific productivity (qP), resisting apoptosis and enhancing oxidative metabolism. The authors previously demonstrated that over-expressing miR-92a results in increases in qP and titer of CHO-IgG cells. However, the mechanisms by which miR-92a enhances qP in CHO cells are still uninvestigated. Here, the authors report the identification of insig1, a regulator of cholesterol biosynthesis, as a target of miR-92a using computational prediction. Both transient and stable over-expression of miR-92a decreased the expression levels of insig1. Insig1 was further validated as a target of miR-92a using 3' UTR reporter assay. Intracellular cholesterol concentration of two high-producing miR-92a clones were significantly increased by ≈30% compared to the blank-transfected pool. Relative Golgi surface area was also found to be 18-26% higher in these clones. Our findings suggest that miR-92a may affect cholesterol metabolism by repressing insig1, resulting in raised intracellular cholesterol levels and Golgi volume and hence enhanced protein secretion. Copyright © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Methotrexate in Atherogenesis and Cholesterol Metabolism

    Directory of Open Access Journals (Sweden)

    Eric Coomes

    2011-01-01

    Full Text Available Methotrexate is a disease-modifying antirheumatic drug commonly used to treat inflammatory conditions such as rheumatoid arthritis which itself is linked to increased cardiovascular risk. Treatments that target inflammation may also impact the cardiovascular system. While methotrexate improves cardiovascular risk, inhibition of the cyclooxygenase (COX-2 enzyme promotes atherosclerosis. These opposing cardiovascular influences may arise from differing effects on the expression of proteins involved in cholesterol homeostasis. These proteins, ATP-binding cassette transporter (ABC A1 and cholesterol 27-hydroxylase, facilitate cellular cholesterol efflux and defend against cholesterol overload. Methotrexate upregulates expression of cholesterol 27-hydroxylase and ABCA1 via adenosine release, while COX-2 inhibition downregulates these proteins. Adenosine, acting through the A2A and A3 receptors, may upregulate proteins involved in reverse cholesterol transport by cAMP-PKA-CREB activation and STAT inhibition, respectively. Elucidating underlying cardiovascular mechanisms of these drugs provides a framework for developing novel cardioprotective anti-inflammatory medications, such as selective A2A receptor agonists.

  10. ABC Transport Proteins in Cardiovascular Disease-A Brief Summary.

    Science.gov (United States)

    Schumacher, Toni; Benndorf, Ralf A

    2017-04-06

    Adenosine triphosphate (ATP)-binding cassette (ABC) transporters may play an important role in the pathogenesis of atherosclerotic vascular diseases due to their involvement in cholesterol homeostasis, blood pressure regulation, endothelial function, vascular inflammation, as well as platelet production and aggregation. In this regard, ABC transporters, such as ABCA1, ABCG5 and ABCG8, were initially found to be responsible for genetically-inherited syndromes like Tangier diseases and sitosterolemia. These findings led to the understanding of those transporter's function in cellular cholesterol efflux and thereby also linked them to atherosclerosis and cardiovascular diseases (CVD). Subsequently, further ABC transporters, i.e., ABCG1, ABCG4, ABCB6, ABCC1, ABCC6 or ABCC9, have been shown to directly or indirectly affect cellular cholesterol efflux, the inflammatory response in macrophages, megakaryocyte proliferation and thrombus formation, as well as vascular function and blood pressure, and may thereby contribute to the pathogenesis of CVD and its complications. Furthermore, ABC transporters, such as ABCB1, ABCC2 or ABCG2, may affect the safety and efficacy of several drug classes currently in use for CVD treatment. This review will give a brief overview of ABC transporters involved in the process of atherogenesis and CVD pathology. It also aims to briefly summarize the role of ABC transporters in the pharmacokinetics and disposition of drugs frequently used to treat CVD and CVD-related complications.

  11. Differential mRNA expression of seven genes involved in cholesterol metabolism and transport in the liver of atherosclerosis-susceptible and -resistant Japanese quail strains

    Directory of Open Access Journals (Sweden)

    Li Xinrui

    2012-06-01

    Full Text Available Abstract Background Two atherosclerosis-susceptible and -resistant Japanese quail (Coturnix japonica strains obtained by divergent selection are commonly used as models to study atherosclerosis, but no genetic characterization of their phenotypic differences has been reported so far. Our objective was to examine possible differences in the expression of genes involved in cholesterol metabolism and transport in the liver between these two strains and to evaluate the value of this model to analyze the gene system affecting cholesterol metabolism and transport. Methods A factorial study with both strains (atherosclerosis-susceptible versus atherosclerosis-resistant and two diets (control versus cholesterol was carried out. The mRNA concentrations of four genes involved in cholesterol biosynthesis (HMGCR, FDFT1, SQLE and DHCR7 and three genes in cholesterol transport (ABCG5, ABCG8 and APOA1 were assayed using real-time quantitative PCR. Plasma lipids were also assayed. Results Expression of ABCG5 (control diet and ABCG8 (regardless of dietary treatment and expression of HMGCR, FDFT1 and SQLE (regardless of dietary treatment were significantly higher in the atherosclerosis-resistant than in the atherosclerosis-susceptible strain. Plasma triglyceride and LDL levels, and LDL/HDL ratio were significantly higher in the atherosclerosis-susceptible than in the atherosclerosis-resistant strain fed the cholesterol diet. In the atherosclerosis-susceptible strain, ABCG5 expression regressed significantly and positively on plasma LDL level, whereas DHCR7 and SQLE expression regressed significantly and negatively on plasma triglyceride level. Conclusions Our results provide support for the hypothesis that the atherosclerosis-resistant strain metabolizes and excretes cholesterol faster than the atherosclerosis-susceptible strain. We have also demonstrated that these quail strains are a useful model to study cholesterol metabolism and transport in relation with

  12. What's Cholesterol?

    Science.gov (United States)

    ... LDL. Most cholesterol is LDL (low-density lipoprotein) cholesterol. LDL cholesterol is more likely to clog blood vessels because ... Here's a way to remember the difference: the LDL cholesterol is the bad kind, so call it "lousy" ...

  13. Cholesterol and polyunsaturated lipids working in concert to modulate G protein-coupled receptors

    Czech Academy of Sciences Publication Activity Database

    Javanainen, M.; Manna, M.; Vähäheikkilä, M.; Niemelä, M.; Tynkkynen, J.; Guixa-Gonzaléz, R.; Martinez-Seara, Hector; Enkavi, G.; Kulig, W.; Müller, D. J.; Rog, T.; Vattulainen, I.

    2017-01-01

    Roč. 46, Suppl 1 (2017), S47 ISSN 0175-7571. [IUPAB congress /19./ and EBSA congress /11./. 16.07.2017-20.07.2017, Edinburgh] Institutional support: RVO:61388963 Keywords : GPCR * cholesterol * lipids Subject RIV: BO - Biophysics

  14. Cholesterol sensitivity of detergent resistance: A rapid flow cytometric test for detecting constitutive or induced raft association of membrane proteins

    OpenAIRE

    Imre Gombos, Zsolt Bacsó, Cynthia Detre, Henrietta Nagy, Katalin Goda, Márton Andrásfalvy, Gábor Szabó, János Matkó; Bacsó Zsolt (1963-) (biofizikus); Goda Katalin (1969-) (biofizikus); Szabó Gábor (1953-) (biofizikus)

    2004-01-01

    Lipid rafts are cholesterol- and glycosphingolipid-rich microdomains in the cellular plasma membranes that play critical roles in compartmentalization (concentration, coupling, and isolation) of receptors and signal molecules. Therefore, detecting constitutive or induced raft associations of such proteins is of central interest in cell biology. This has mostly been done with time- and cell-consuming immunobiochemical techniques affected by several sources of artifacts. A flow cytometric analy...

  15. Actin binding proteins, spermatid transport and spermiation*

    Science.gov (United States)

    Qian, Xiaojing; Mruk, Dolores D.; Cheng, Yan-Ho; Tang, Elizabeth I.; Han, Daishu; Lee, Will M.; Wong, Elissa W. P.; Cheng, C. Yan

    2014-01-01

    The transport of germ cells across the seminiferous epithelium is composed of a series of cellular events during the epithelial cycle essential to the completion of spermatogenesis. Without the timely transport of spermatids during spermiogenesis, spermatozoa that are transformed from step 19 spermatids in the rat testis fail to reach the luminal edge of the apical compartment and enter the tubule lumen at spermiation, thereby entering the epididymis for further maturation. Step 19 spermatids and/or sperms that remain in the epithelium will be removed by the Sertoli cell via phagocytosis to form phagosomes and be degraded by lysosomes, leading to subfertility and/or infertility. However, the biology of spermatid transport, in particular the final events that lead to spermiation remain elusive. Based on recent data in the field, we critically evaluate the biology of spermiation herein by focusing on the actin binding proteins (ABPs) that regulate the organization of actin microfilaments at the Sertoli-spermatid interface, which is crucial for spermatid transport during this event. The hypothesis we put forth herein also highlights some specific areas of research that can be pursued by investigators in the years to come. PMID:24735648

  16. Distribution and Spectroscopy of Green Fluorescent Protein and Acyl-CoA: Cholesterol Acytransferase in Sf21 Insect Cells

    Science.gov (United States)

    Richmond, R. C.; Mahtani, H.; Lu, X.; Chang, T. Y.; Malak, H.; Rose, M. Franklin (Technical Monitor)

    2001-01-01

    Acyl-CoA: cholesterol acyltransferase (ACAT) is thought to significantly participate in the pathway of cholesterol esterification that underlies the pathology of artherosclerosis. This enzyme is a membrane protein known to be preferentially bound within the endoplasmic reticulum of mammalian cells, from which location it esterifies cholesterol derived from low density lipoprotein. Cultures of insect cells were separately infected with baculovirus containing the gene for green fluroescent protein (GFP) and with baculovirus containing tandem genes for GFP and ACAT. These infected cultures expressed GFP and the fusion protein GCAT, respectively, with maximum expression occurring on the fourth day after infection. Extraction of GFP- and of GCAT-expressing cells with urea and detergent resulted in recovery of fluorescent protein in aqueous solution. Fluorescence spectra at neutral pH were identical for both GFP and GCAT extracts in aqueous solution, indicating unperturbed tertiary structure for the GFP moiety within GCAT. In a cholesterol esterification assay, GCAT demonstrated ACAT activity, but with less efficiency compared to native ACAT. It was hypothesized that the membrane protein ACAT would lead to differences in localization of GCAT compared to GFP within the respective expressing insect cells. The GFP marker directly and also within the fusion protein GCAT was accordingly used as the intracellular probe that was fluorescently analyzed by the new biophotonics technique of hyperspectral imaging. In that technique, fluorescence imaging was obtained from two dimensional arrays of cells, and regions of interest from within those images were then retrospectively analyzed for the emission spectra that comprises the image. Results of hyperspectral imaging of insect cells on day 4 postinfection showed that GCAT was preferentially localized to the cytoplasm of these cells compared to GFP. Furthermore, the emission spectra obtained for the localized GCAT displayed a peak

  17. Complement activation by ceramide transporter proteins.

    Science.gov (United States)

    Bode, Gerard H; Losen, Mario; Buurman, Wim A; Veerhuis, Robert; Molenaar, Peter C; Steinbusch, Harry W M; De Baets, Marc H; Daha, Mohamed R; Martinez-Martinez, Pilar

    2014-02-01

    C1q is the initiator of the classical complement pathway and, as such, is essential for efficient opsonization and clearance of pathogens, altered self-structures, and apoptotic cells. The ceramide transporter protein (CERT) and its longer splicing isoform CERTL are known to interact with extracellular matrix components, such as type IV collagen, and with the innate immune protein serum amyloid P. In this article, we report a novel function of CERT in the innate immune response. Both CERT isoforms, when immobilized, were found to bind the globular head region of C1q and to initiate the classical complement pathway, leading to activation of C4 and C3, as well as generation of the membrane attack complex C5b-9. In addition, C1q was shown to bind to endogenous CERTL on the surface of apoptotic cells. These results demonstrate the role of CERTs in innate immunity, especially in the clearance of apoptotic cells.

  18. Rosuvastatin activates ATP-binding cassette transporter A1-dependent efflux ex vivo and promotes reverse cholesterol transport in macrophage cells in mice fed a high-fat diet.

    Science.gov (United States)

    Shimizu, Tomohiko; Miura, Shin-ichiro; Tanigawa, Hiroyuki; Kuwano, Takashi; Zhang, Bo; Uehara, Yoshinari; Saku, Keijiro

    2014-10-01

    It is controversial whether statins improve high-density lipoprotein (HDL) function, which plays an important role in reverse cholesterol transport in vivo. The aim of the present study was to clarify the effects of rosuvastatin and atorvastatin on reverse cholesterol transport in macrophage cells in vivo and their underlying mechanisms. Male C57BL mice were divided into 3 groups (rosuvastatin, atorvastatin, and control groups) and orally administered rosuvastatin, atorvastatin, or placebo for 6 weeks under feeding with a 0.5% cholesterol+10% coconut oil diet. After administration, although there were no changes in plasma HDL cholesterol levels among the groups, plasma from the rosuvastatin group showed an increased ability to promote ATP-binding cassette transporter A1-mediated cholesterol efflux ex vivo. In addition, capillary electrophoresis revealed a shift in HDL toward the pre-β HDL fraction only in the rosuvastatin group. Mice in all 3 groups were intraperitoneally injected with (3)H-cholesterol-labeled and cholesterol-loaded macrophages and then were monitored for the appearance of (3)H-tracer in plasma and feces. The amount of (3)H-tracer excreted into feces during 48 hours in the rosuvastatin group was greater than that in the control group. Finally, (3)H-cholesteryl oleate-HDL was intravenously injected into all groups, blood samples were taken, and the count of (3)H-cholesterol was analyzed. Plasma (3)H-cholesteryl oleate-HDL changed similarly, and no differences in fractional catabolic rates were observed. Rosuvastatin enhanced the ATP-binding cassette transporter A1-dependent HDL efflux function of reverse cholesterol transport, and this finding highlights the potential of rosuvastatin for the regression of atherosclerosis. © 2014 American Heart Association, Inc.

  19. Isoflavone and protein constituents of lactic acid-fermented soy milk combine to prevent dyslipidemia in rats fed a high cholesterol diet.

    Science.gov (United States)

    Kobayashi, Maki; Egusa, Shintaro; Fukuda, Mitsuru

    2014-12-10

    A high cholesterol diet induces dyslipidemia. This study investigated whether isoflavone aglycones in lactic acid-fermented soy milk (LFS) improve lipid metabolism in rats fed a high cholesterol diet. Male Sprague-Dawley rats aged seven weeks were fed an AIN-93G diet, a 1% cholesterol diet (a high cholesterol diet), a high-cholesterol diet containing 4% isoflavone extract of LFS (LFS extract diet), a high-cholesterol diet containing 19.4% ethanol-washed LFS (ethanol-washed LFS diet, isoflavone-poor diet), or a high cholesterol diet containing 23.2% intact LFS (intact LFS diet) for five weeks. The plasma total cholesterol (TC) level was increased in the rats fed the LFS extract diet compared with those fed the high cholesterol diet. The TC level was decreased by the intact LFS and ethanol-washed LFS diets. The cholesterol-lowering effect was stronger in the rats fed the intact LFS diet than those fed the ethanol-washed LFS diet. The plasma triglyceride (TG) level was unchanged in the rats fed the LFS extract diet, but it decreased in rats fed the intact LFS and ethanol-washed LFS diets. Although, compared with the high cholesterol diet, the LFS extract and ethanol-washed LFS diets did not reduce hepatic cholesterol and TG, both levels were remarkably lowered by the intact LFS diet. These results suggest that the improvement in lipid metabolism of rats fed a high-cholesterol diet containing LFS isoflavone aglycones is not due to an independent effect but due to a cooperative effect with soy protein.

  20. Isoflavone and Protein Constituents of Lactic Acid-Fermented Soy Milk Combine to Prevent Dyslipidemia in Rats Fed a High Cholesterol Diet

    Directory of Open Access Journals (Sweden)

    Maki Kobayashi

    2014-12-01

    Full Text Available A high cholesterol diet induces dyslipidemia. This study investigated whether isoflavone aglycones in lactic acid-fermented soy milk (LFS improve lipid metabolism in rats fed a high cholesterol diet. Male Sprague-Dawley rats aged seven weeks were fed an AIN-93G diet, a 1% cholesterol diet (a high cholesterol diet, a high-cholesterol diet containing 4% isoflavone extract of LFS (LFS extract diet, a high-cholesterol diet containing 19.4% ethanol-washed LFS (ethanol-washed LFS diet, isoflavone-poor diet, or a high cholesterol diet containing 23.2% intact LFS (intact LFS diet for five weeks. The plasma total cholesterol (TC level was increased in the rats fed the LFS extract diet compared with those fed the high cholesterol diet. The TC level was decreased by the intact LFS and ethanol-washed LFS diets. The cholesterol-lowering effect was stronger in the rats fed the intact LFS diet than those fed the ethanol-washed LFS diet. The plasma triglyceride (TG level was unchanged in the rats fed the LFS extract diet, but it decreased in rats fed the intact LFS and ethanol-washed LFS diets. Although, compared with the high cholesterol diet, the LFS extract and ethanol-washed LFS diets did not reduce hepatic cholesterol and TG, both levels were remarkably lowered by the intact LFS diet. These results suggest that the improvement in lipid metabolism of rats fed a high-cholesterol diet containing LFS isoflavone aglycones is not due to an independent effect but due to a cooperative effect with soy protein.

  1. EhNPC1 and EhNPC2 Proteins Participate in Trafficking of Exogenous Cholesterol in Entamoeba histolytica Trophozoites: Relevance for Phagocytosis.

    Directory of Open Access Journals (Sweden)

    Jeni Bolaños

    2016-12-01

    Full Text Available Entamoeba histolytica, the highly phagocytic protozoan causative of human amoebiasis lacks the machinery to synthesize cholesterol. Here, we investigated the presence of NPC1 and NPC2 proteins in this parasite, which are involved in cholesterol trafficking in mammals. Bioinformatics analysis revealed one Ehnpc1 and two Ehnpc2 genes. EhNPC1 appeared as a transmembrane protein and both EhNPC2 as peripheral membrane proteins. Molecular docking predicted that EhNPC1 and EhNPC2 bind cholesterol and interact with each other. Genes and proteins were identified in trophozoites. Serum pulse-chase and confocal microscopy assays unveiled that after trophozoites sensed the cholesterol source, EhNPC1 and EhNPC2 were organized around the plasma membrane in a punctuated pattern. Vesicles emerged and increased in number and size and some appeared full of cholesterol with EhNPC1 or EhNPC2 facing the extracellular space. Both proteins, but mostly EhNPC2, were found out of the cell associated with cholesterol. EhNPC1 and cholesterol formed networks from the plasma membrane to the nucleus. EhNPC2 appeared in erythrocytes that were being ingested by trophozoites, co-localizing with cholesterol of erythrocytes, whereas EhNPC1 surrounded the phagocytic cup. EhNPC1 and EhNPC2 co-localized with EhSERCA in the endoplasmic reticulum and with lysobisphosphatidic acid and EhADH (an Alix protein in phagolysosomes. Immunoprecipitation assays confirmed the EhNPC1 and EhNPC2 association with cholesterol, EhRab7A and EhADH. Serum starved and blockage of cholesterol trafficking caused a low rate of phagocytosis and incapability of trophozoites to produce damage in the mouse colon. Ehnpc1 and Ehnpc2 knockdown provoked in trophozoites a lower intracellular cholesterol concentration and a diminished rate of phagocytosis; and Ehnpc1 silencing also produced a decrease of trophozoites movement. Trafficking of EhNPC1 and EhNPC2 during cholesterol uptake and phagocytosis as well as their

  2. Increased cholesterol efflux from cultured fibroblasts to plasma from hypertriglyceridemic type 2 diabetic patients : Roles of pre beta-HDL, phospholipid transfer protein and cholesterol esterification

    NARCIS (Netherlands)

    de Vries, R.; Groen, A. K.; Perton, F. G.; Dallinga-Thie, G. M.; van Wijland, M. J. A.; Dikkeschei, L. D.; Wolffenbuttel, B. H. R.; van Tol, A.; Dullaart, R. P. F.

    We tested whether hypertriglyceridemia associated with type 2 diabetes mellitus is accompanied by alterations in pre beta-HDL, which are considered to be initial acceptors of cell-derived cholesterol, and by changes in the ability of plasma to promote cellular cholesterol efflux. In 28

  3. MicroRNA-19b promotes macrophage cholesterol accumulation and aortic atherosclerosis by targeting ATP-binding cassette transporter A1.

    Science.gov (United States)

    Lv, Yun-Cheng; Tang, Yan-Yan; Peng, Juan; Zhao, Guo-Jun; Yang, Jing; Yao, Feng; Ouyang, Xin-Ping; He, Ping-Ping; Xie, Wei; Tan, Yu-Lin; Zhang, Min; Liu, Dan; Tang, Deng-Pei; Cayabyab, Francisco S; Zheng, Xi-Long; Zhang, Da-Wei; Tian, Guo-Ping; Tang, Chao-Ke

    2014-09-01

    Macrophage accumulation of cholesterol leads to foam cell formation which is a major pathological event of atherosclerosis. Recent studies have shown that microRNA (miR)-19b might play an important role in cholesterol metabolism and atherosclerotic diseases. Here, we have identified miR-19b binding to the 3'UTR of ATP-binding cassette transporter A1 (ABCA1) transporters, and further determined the potential roles of this novel interaction in atherogenesis. To investigate the molecular mechanisms involved in a miR-19b promotion of macrophage cholesterol accumulation and the development of aortic atherosclerosis. We performed bioinformatics analysis using online websites, and found that miR-19b was highly conserved during evolution and directly bound to ABCA1 mRNA with very low binding free energy. Luciferase reporter assay confirmed that miR-19b bound to 3110-3116 sites within ABCA1 3'UTR. MiR-19b directly regulated the expression levels of endogenous ABCA1 in foam cells derived from human THP-1 macrophages and mouse peritoneal macrophages (MPMs) as determined by qRT-PCR and western blot. Cholesterol transport assays revealed that miR-19b dramatically suppressed apolipoprotein AI-mediated ABCA1-dependent cholesterol efflux, resulting in the increased levels of total cholesterol (TC), free cholesterol (FC) and cholesterol ester (CE) as revealed by HPLC. The excretion of (3)H-cholesterol originating from cholesterol-laden MPMs into feces was decreased in mice overexpressing miR-19b. Finally, we evaluated the proatherosclerotic role of miR-19b in apolipoprotein E deficient (apoE(-/-)) mice. Treatment with miR-19b precursor reduced plasma high-density lipoprotein (HDL) levels, but increased plasma low-density lipoprotein (LDL) levels. Consistently, miR-19b precursor treatment increased aortic plaque size and lipid content, but reduced collagen content and ABCA1 expression. In contrast, treatment with the inhibitory miR-19b antisense oligonucleotides (ASO) prevented or

  4. Simvastatin reduces atherogenesis and promotes the expression of hepatic genes associated with reverse cholesterol transport in apoE-knockout mice fed high-fat diet

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    Tian Hua

    2011-01-01

    Full Text Available Abstract Background Statins are first-line pharmacotherapeutic agents for hypercholesterolemia treatment in humans. However the effects of statins on atherosclerosis in mouse models are very paradoxical. In this work, we wanted to evaluate the effects of simvastatin on serum cholesterol, atherogenesis, and the expression of several factors playing important roles in reverse cholesterol transport (RCT in apoE-/- mice fed a high-fat diet. Results The atherosclerotic lesion formation displayed by oil red O staining positive area was reduced significantly by 35% or 47% in either aortic root section or aortic arch en face in simvastatin administrated apoE-/- mice compared to the control. Plasma analysis by enzymatic method or ELISA showed that high-density lipoprotein-cholesterol (HDL-C and apolipoprotein A-I (apoA-I contents were remarkably increased by treatment with simvastatin. And plasma lecithin-cholesterol acyltransferase (LCAT activity was markedly increased by simvastatin treatment. Real-time PCR detection disclosed that the expression of several transporters involved in reverse cholesterol transport, including macrophage scavenger receptor class B type I, hepatic ATP-binding cassette (ABC transporters ABCG5, and ABCB4 were induced by simvastatin treatment, the expression of hepatic ABCA1 and apoA-I, which play roles in the maturation of HDL-C, were also elevated in simvastatin treated groups. Conclusions We demonstrated the anti-atherogenesis effects of simvastatin in apoE-/- mice fed a high-fat diet. We confirmed here for the first time simvastatin increased the expression of hepatic ABCB4 and ABCG5, which involved in secretion of cholesterol and bile acids into the bile, besides upregulated ABCA1 and apoA-I. The elevated HDL-C level, increased LCAT activity and the stimulation of several transporters involved in RCT may all contribute to the anti-atherosclerotic effect of simvastatin.

  5. Two-compartment model as a teaching tool for cholesterol homeostasis.

    Science.gov (United States)

    Wrona, Artur; Balbus, Joanna; Hrydziuszko, Olga; Kubica, Krystian

    2015-12-01

    Cholesterol is a vital structural and functional molecule in the human body that is only slightly soluble in water and therefore does not easily travels by itself in the bloodstream. To enable cholesterol's targeted delivery to cells and tissues, it is encapsulated by different fractions of lipoproteins, complex particles containing both proteins and lipids. Maintaining cholesterol homeostasis is a highly regulated process with multiple factors acting at both molecular and tissue levels. Furthermore, to regulate the circulatory transport of cholesterol in lipoproteins, the amount of cholesterol present depends on and is controlled by cholesterol dietary intake, de novo synthesis, usage, and excretion; abnormal and/or unbalanced cholesterol levels have been shown to lead to severe outcomes, e.g., cardiovascular diseases. To investigate cholesterol transport in the circulatory system, we have previously developed a two-compartment mathematical model. Here, we show how this model can be used as a teaching tool for cholesterol homeostasis. Using the model and a hands-on approach, students can familiarize themselves with the basic components and mechanisms behind balanced cholesterol circulatory transport as well as investigate the consequences of and countermeasures to abnormal cholesterol levels. Among others, various treatments of high blood cholesterol levels can be simulated, e.g., with commonly prescribed de novo cholesterol synthesis inhibitors. Copyright © 2015 The American Physiological Society.

  6. Cholesterol Removal from Adult Skeletal Muscle impairs Excitation-Contraction Coupling and Aging reduces Caveolin-3 and alters the Expression of other Triadic Proteins

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    Genaro eBarrientos

    2015-04-01

    Full Text Available Cholesterol and caveolin are integral membrane components that modulate the function/location of many cellular proteins. Skeletal muscle fibers, which have unusually high cholesterol levels in transverse tubules, express the caveolin-3 isoform but its association with transverse tubules remains contentious. Cholesterol removal impairs excitation-contraction coupling in amphibian and mammalian fetal skeletal muscle fibers. Here, we show that treating single muscle fibers from adult mice with the cholesterol removing agent methyl-β-cyclodextrin decreased fiber cholesterol by 26%, altered the location pattern of caveolin-3 and of the voltage dependent calcium channel Cav1.1, and suppressed or reduced electrically evoked Ca2+ transients without affecting membrane integrity or causing sarcoplasmic reticulum calcium depletion. We found that transverse tubules from adult muscle and triad fractions that contain ~10% attached transverse tubules, but not sarcoplasmic reticulum membranes, contained caveolin-3 and Cav1.1; both proteins partitioned into detergent-resistant membrane fractions highly enriched in cholesterol. Aging entails significant deterioration of skeletal muscle function. We found that triad fractions from aged rats had similar cholesterol and RyR1 protein levels compared to triads from young rats, but had lower caveolin-3 and glyceraldehyde 3-phosphate dehydrogenase and increased Na+/K+-ATPase protein levels. Both triad fractions had comparable NADPH oxidase (NOX activity and protein content of NOX2 subunits (p47phox and gp91phox, implying that NOX activity does not increase during aging. These findings show that partial cholesterol removal impairs excitation-contraction coupling and alters caveolin-3 and Cav1.1 location pattern, and that aging reduces caveolin-3 protein content and modifies the expression of other triadic proteins. We discuss the possible implications of these findings for skeletal muscle function in young and aged

  7. MicroRNA-20a/b regulates cholesterol efflux through post-transcriptional repression of ATP-binding cassette transporter A1.

    Science.gov (United States)

    Liang, Bin; Wang, Xin; Song, Xiaosu; Bai, Rui; Yang, Huiyu; Yang, Zhiming; Xiao, Chuanshi; Bian, Yunfei

    2017-09-01

    ATP-binding cassette transporter A1 (ABCA1) plays a crucial role in reverse cholesterol transport and exhibits anti-atherosclerosis effects. Some microRNAs (miRs) regulate ABCA1 expression, and recent studies have shown that miR-20a/b might play a critical role in atherosclerotic diseases. Here, we attempted to clarify the potential contribution of miR-20a/b in post-transcriptional regulation of ABCA1, cholesterol efflux, and atherosclerosis. We performed bioinformatics analysis and found that miR-20a/b was highly conserved and directly bound to ABCA1 mRNA with low binding free energy. Luciferase-reporter assay also confirmed that miR-20a/b significantly reduced luciferase activity associated with the ABCA1 3' untranslated region reporter construct. Additionally, miR-20a/b decreased ABCA1 expression, which, in turn, decreased cholesterol efflux and increased cholesterol content in THP-1 and RAW 264.7 macrophage-derived foam cells. In contrast, miR-20a/b inhibitors increased ABCA1 expression and cholesterol efflux, decreased cholesterol content, and inhibited foam-cell formation. Consistent with our in vitro results, miR-20a/b-treated ApoE -/- mice showed decreased ABCA1expression in the liver and reductions of reverse cholesterol transport in vivo. Furthermore, miR-20a/b regulated the formation of nascent high-density lipoprotein and promoted atherosclerotic development, whereas miR-20a/b knockdown attenuated atherosclerotic formation. miR-20 is a new miRNA capable of targeting ABCA1 and regulating ABCA1 expression. Therefore, miR-20 inhibition constitutes a new strategy for ABCA1-based treatment of atherosclerosis. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. An Essential Role of Hrs/Vps27 in Endosomal Cholesterol Trafficking

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    Ximing Du

    2012-01-01

    Full Text Available The endosomal sorting complex required for transport (ESCRT plays a crucial role in the degradation of ubiquitinated endosomal membrane proteins. Here, we report that Hrs, a key protein of the ESCRT-0 complex, is required for the transport of low-density lipoprotein-derived cholesterol from endosomes to the endoplasmic reticulum. This function of Hrs in cholesterol transport is distinct from its previously defined role in lysosomal sorting and downregulation of membrane receptors via the ESCRT pathway. In line with this, knocking down other ESCRT proteins does not cause prominent endosomal cholesterol accumulation. Importantly, the localization and biochemical properties of key cholesterol-sorting proteins, NPC1 and NPC2, appear to be unchanged upon Hrs knockdown. Our data identify Hrs as a regulator of endosomal cholesterol trafficking and provide additional insights into the budding of intralumenal vesicles.

  9. Cholesterol through the Looking Glass

    Science.gov (United States)

    Kristiana, Ika; Luu, Winnie; Stevenson, Julian; Cartland, Sian; Jessup, Wendy; Belani, Jitendra D.; Rychnovsky, Scott D.; Brown, Andrew J.

    2012-01-01

    How cholesterol is sensed to maintain homeostasis has been explained by direct binding to a specific protein, Scap, or through altering the physical properties of the membrane. The enantiomer of cholesterol (ent-cholesterol) is a valuable tool in distinguishing between these two models because it shares nonspecific membrane effects with native cholesterol (nat-cholesterol), but not specific binding interactions. This is the first study to compare ent- and nat-cholesterol directly on major molecular parameters of cholesterol homeostasis. We found that ent-cholesterol suppressed activation of the master transcriptional regulator of cholesterol metabolism, SREBP-2, almost as effectively as nat-cholesterol. Importantly, ent-cholesterol induced a conformational change in the cholesterol-sensing protein Scap in isolated membranes in vitro, even when steps were taken to eliminate potential confounding effects from endogenous cholesterol. Ent-cholesterol also accelerated proteasomal degradation of the key cholesterol biosynthetic enzyme, squalene monooxygenase. Together, these findings provide compelling evidence that cholesterol maintains its own homeostasis not only via direct protein interactions, but also by altering membrane properties. PMID:22869373

  10. Tritium Suicide Selection Identifies Proteins Involved in the Uptake and Intracellular Transport of Sterols in Saccharomyces cerevisiae▿

    Science.gov (United States)

    Sullivan, David P.; Georgiev, Alexander; Menon, Anant K.

    2009-01-01

    Sterol transport between the plasma membrane (PM) and the endoplasmic reticulum (ER) occurs by a nonvesicular mechanism that is poorly understood. To identify proteins required for this process, we isolated Saccharomyces cerevisiae mutants with defects in sterol transport. We used Upc2-1 cells that have the ability to take up sterols under aerobic conditions and exploited the observation that intracellular accumulation of exogenously supplied [3H]cholesterol in the form of [3H]cholesteryl ester requires an intact PM-ER sterol transport pathway. Upc2-1 cells were mutagenized using a transposon library, incubated with [3H]cholesterol, and subjected to tritium suicide selection to isolate mutants with a decreased ability to accumulate [3H]cholesterol. Many of the mutants had defects in the expression and trafficking of Aus1 and Pdr11, PM-localized ABC transporters that are required for sterol uptake. Through characterization of one of the mutants, a new role was uncovered for the transcription factor Mot3 in controlling expression of Aus1 and Pdr11. A number of mutants had transposon insertions in the uncharacterized Ydr051c gene, which we now refer to as DET1 (decreased ergosterol transport). These mutants expressed Aus1 and Pdr11 normally but were severely defective in the ability to accumulate exogenously supplied cholesterol. The transport of newly synthesized sterols from the ER to the PM was also defective in det1Δ cells. These data indicate that the cytoplasmic protein encoded by DET1 is involved in intracellular sterol transport. PMID:19060182

  11. Detergent insolubility of alkaline phosphatase during biosynthetic transport and endocytosis. Role of cholesterol

    NARCIS (Netherlands)

    Cerneus, D. P.; Ueffing, E.; Posthuma, G.; Strous, G. J.; van der Ende, A.

    1993-01-01

    Alkaline phosphatase is anchored to the outer leaflet of the plasma membrane by a covalently attached glycosyl-phosphatidylinositol anchor. We have studied the biosynthetic transport and endocytosis of alkaline phosphatase in the choriocarcinoma cell line BeWo, which endogenously expresses this

  12. A new cytolytic protein from the sea anemone Urticina crassicornis that binds to cholesterol- and sphingomyelin-rich membranes.

    Science.gov (United States)

    Razpotnik, Andrej; Krizaj, Igor; Kem, William R; Macek, Peter; Turk, Tom

    2009-06-01

    A new pore-forming cytolytic protein was isolated from the Northern red sea anemone, Urticina crassicornis. Its biochemical properties were characterized and partial N-terminal amino acid sequence was determined. The cytolysin, named UcI, has a molecular mass of around 30kDa and lacks phospholipase A(2) activity. UcI lyses bovine erythrocytes at nanomolar concentrations. Hemolysis is a result of a colloid-osmotic shock caused by the opening of toxin-induced ionic pores and can be prevented by osmotic protectants of size >600Da. The functional radius of an average pore was estimated to be about 0.66nm. A more detailed study of the cytolytic activity of UcI was performed with lipid vesicles and monolayers. The toxin binds to monolayers and efficiently permeabilizes small lipid vesicles composed of sphingomyelin and cholesterol. However, the cytolytic activity is not prevented by preincubation with either pure cholesterol or sphingomyelin dispersions. We conclude that the presence of both sphingomyelin and cholesterol, key components of lipid rafts, greatly enhances toxin binding to membranes and probably facilitates pore formation. Alignment of the toxin partial amino acid sequence with sequences of cytolysins belonging to the actinoporin family reveals no sequence homology. We conclude that partial sequence of UcI resembles only the N-terminal part of UpI, a cytolytic protein isolated from a related sea anemone species, Urticina piscivora. The two proteins most probably belong to a separate family of sea anemone cytolysins that are worthy of further characterization.

  13. High pressure modulated transport and signaling functions of membrane proteins in models and in vivo

    International Nuclear Information System (INIS)

    Vogel, R F; Linke, K; Teichert, H; Ehrmann, M A

    2008-01-01

    Cellular membranes serve in the separation of compartments, recognition of the environment, selective transport and signal transduction. Membrane lipids and membrane proteins play distinct roles in these processes, which are affected by environmental chemical (e. g. pH) or physical (e. g. pressure and temperature) changes. High hydrostatic pressure (HHP) affects fluidity and integrity of bacterial membranes instantly during the ramp, resulting in a loss of membrane potential and vital membrane protein functions. We have used the multiple drug transporter LmrA from Lactococcus lactis and ToxR, a membrane protein sensor from Photobacterium profundum, a deep-sea bacterium, and Vibrio cholerae to study membrane protein interaction and functionality in proteolioposomes and by the use of in vivo reporter systems, respectively. Both proteins require dimerization in the phospholipid bilayer for their functionality, which was favoured in the liquid crystalline lipid phase with ToxR and LmrA. Whereas LmrA, which resides in liposomes consisting of DMPC, DMPC/cholesterol or natural lipids, lost its ATPase activity above 20 or 40 MPa, it maintained its active dimeric structure in DOPC/DPPC/cholesterol liposomes up to 120 MPa. By using a specific indicator strain in which the dimerisation of ToxR initiates the transcription of lacZ it was demonstrated, that the amino acid sequence of the transmembrane domain influences HHP stability of ToxR dimerization in vivo. Thus, both the lipid structure and the nature of the protein affect membrane protein interaction. It is suggested that the protein structure determines basic functionality, e.g. principle ability or kinetics to dimerize to a functional complex, while the lipid environment modulates this property

  14. High pressure modulated transport and signaling functions of membrane proteins in models and in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Vogel, R F; Linke, K; Teichert, H; Ehrmann, M A [Technische Universitaet Muenchen, Technische Mikrobiologie, Weihenstephaner Steig 16, 85350 Freising (Germany)], E-mail: rudi.vogel@wzw.tum.de

    2008-07-15

    Cellular membranes serve in the separation of compartments, recognition of the environment, selective transport and signal transduction. Membrane lipids and membrane proteins play distinct roles in these processes, which are affected by environmental chemical (e. g. pH) or physical (e. g. pressure and temperature) changes. High hydrostatic pressure (HHP) affects fluidity and integrity of bacterial membranes instantly during the ramp, resulting in a loss of membrane potential and vital membrane protein functions. We have used the multiple drug transporter LmrA from Lactococcus lactis and ToxR, a membrane protein sensor from Photobacterium profundum, a deep-sea bacterium, and Vibrio cholerae to study membrane protein interaction and functionality in proteolioposomes and by the use of in vivo reporter systems, respectively. Both proteins require dimerization in the phospholipid bilayer for their functionality, which was favoured in the liquid crystalline lipid phase with ToxR and LmrA. Whereas LmrA, which resides in liposomes consisting of DMPC, DMPC/cholesterol or natural lipids, lost its ATPase activity above 20 or 40 MPa, it maintained its active dimeric structure in DOPC/DPPC/cholesterol liposomes up to 120 MPa. By using a specific indicator strain in which the dimerisation of ToxR initiates the transcription of lacZ it was demonstrated, that the amino acid sequence of the transmembrane domain influences HHP stability of ToxR dimerization in vivo. Thus, both the lipid structure and the nature of the protein affect membrane protein interaction. It is suggested that the protein structure determines basic functionality, e.g. principle ability or kinetics to dimerize to a functional complex, while the lipid environment modulates this property.

  15. High pressure modulated transport and signaling functions of membrane proteins in models and in vivo

    Science.gov (United States)

    Vogel, R. F.; Linke, K.; Teichert, H.; Ehrmann, M. A.

    2008-07-01

    Cellular membranes serve in the separation of compartments, recognition of the environment, selective transport and signal transduction. Membrane lipids and membrane proteins play distinct roles in these processes, which are affected by environmental chemical (e. g. pH) or physical (e. g. pressure and temperature) changes. High hydrostatic pressure (HHP) affects fluidity and integrity of bacterial membranes instantly during the ramp, resulting in a loss of membrane potential and vital membrane protein functions. We have used the multiple drug transporter LmrA from Lactococcus lactis and ToxR, a membrane protein sensor from Photobacterium profundum, a deep-sea bacterium, and Vibrio cholerae to study membrane protein interaction and functionality in proteolioposomes and by the use of in vivo reporter systems, respectively. Both proteins require dimerization in the phospholipid bilayer for their functionality, which was favoured in the liquid crystalline lipid phase with ToxR and LmrA. Whereas LmrA, which resides in liposomes consisting of DMPC, DMPC/cholesterol or natural lipids, lost its ATPase activity above 20 or 40 MPa, it maintained its active dimeric structure in DOPC/DPPC/cholesterol liposomes up to 120 MPa. By using a specific indicator strain in which the dimerisation of ToxR initiates the transcription of lacZ it was demonstrated, that the amino acid sequence of the transmembrane domain influences HHP stability of ToxR dimerization in vivo. Thus, both the lipid structure and the nature of the protein affect membrane protein interaction. It is suggested that the protein structure determines basic functionality, e.g. principle ability or kinetics to dimerize to a functional complex, while the lipid environment modulates this property.

  16. Pharmacological activation of LXR in utero directly influences ABC transporter expression and function in mice but does not affect adult cholesterol metabolism

    NARCIS (Netherlands)

    van Straten, E. M. E.; Huijkman, N. C. A.; Baller, J. F. W.; Kuipers, F.; Plosch, T.

    2008-01-01

    van Straten EM, Huijkman NC, Baller JF, Kuipers F, Plosch T. Pharmacological activation of LXR in utero directly influences ABC transporter expression and function in mice but does not affect adult cholesterol metabolism. Am J Physiol Endocrinol Metab 295: E1341-E1348, 2008. First published October

  17. Inhibition of the apical sodium-dependent bile acid transporter reduces LDL cholesterol and apoB by enhanced plasma clearance of LDL apoB.

    Science.gov (United States)

    Huff, Murray W; Telford, Dawn E; Edwards, Jane Y; Burnett, John R; Barrett, P Hugh R; Rapp, Stephen R; Napawan, Nida; Keller, Bradley T

    2002-11-01

    Cloning of the ileal apical sodium-dependent bile acid transporter (ASBT) has identified a new pharmacological target for the modulation of plasma lipoproteins. The objective of this study was to determine whether a novel, specific, minimally absorbed ASBT inhibitor (SC-435) decreases LDL cholesterol through the alteration of plasma apoB kinetics. Miniature pigs were treated for 21 days with 10 mg/kg/day of SC-435 or placebo. SC-435 decreased plasma cholesterol by 9% and LDL cholesterol by 20% with no effect on other lipids. Autologous (131)I-VLDL, (125)I-LDL, and [(3)H]-leucine were injected simultaneously to determine apoB kinetics. LDL apoB concentrations decreased significantly by 10% resulting entirely from an increase in LDL-apoB fractional catabolic rate. SC-435 had no effect on either total LDL apoB production or VLDL apoB converted to LDL. SC-435 increased VLDL apoB production by 22%; however, the concentration was unchanged as a result of increased VLDL apoB direct removal. SC-435 increased hepatic mRNA and enzymatic activity for both cholesterol 7alpha-hydroxylase and HMG-CoA reductase. Hepatic LDL receptor mRNA increased significantly, whereas apoB expression was unaffected. A low dose of the ASBT inhibitor, SC-435, significantly reduces plasma LDL cholesterol through enhanced LDL receptor-mediated LDL apoB clearance, secondary to increased expression of cholesterol 7alpha-hydroxylase.

  18. Fluorescence spectroscopy studies of HEK293 cells expressing DOR-Gi1alfa fusion protein; the effect of cholesterol depletion

    Czech Academy of Sciences Publication Activity Database

    Brejchová, Jana; Sýkora, Jan; Dlouhá, Kateřina; Roubalová, Lenka; Ostašov, Pavel; Vošahlíková, Miroslava; Hof, Martin; Svoboda, Petr

    2011-01-01

    Roč. 1808, č. 12 (2011), s. 2819-2829 ISSN 0005-2736 R&D Projects: GA MŠk(CZ) LC06063; GA MŠk(CZ) LC554; GA ČR(CZ) GD305/08/H037 Grant - others:GA ČR(CZ) GAP208/10/1090 Institutional research plan: CEZ:AV0Z50110509; CEZ:AV0Z40400503 Keywords : plasma membrane * cholesterol depletion * fluorescence spectroscopy * hydrophobic membrane interior * delta-opioid receptor ( DOR ), * G protein Subject RIV: BO - Biophysics Impact factor: 3.990, year: 2011

  19. Proteins mediating intra- and intercellular transport of lipids and lipid-modified proteins

    NARCIS (Netherlands)

    Neumann, S.

    2008-01-01

    Proteins mediating intra- and intercellular transport of lipids and lipid-modified proteins In this thesis, I studied the intra- and intercellular transport of lipidic molecules, in particular glycosphingolipids and lipid-modified proteins. The first part focuses on the intracellular transport of

  20. Phytosterol and cholesterol precursor levels indicate increased cholesterol excretion and biosynthesis in gallstone disease.

    Science.gov (United States)

    Krawczyk, Marcin; Lütjohann, Dieter; Schirin-Sokhan, Ramin; Villarroel, Luis; Nervi, Flavio; Pimentel, Fernando; Lammert, Frank; Miquel, Juan Francisco

    2012-05-01

    In hepatocytes and enterocytes sterol uptake and secretion is mediated by Niemann-Pick C1-like 1 (NPC1L1) and ATP-binding cassette (ABC)G5/8 proteins, respectively. Whereas serum levels of phytosterols represent surrogate markers for intestinal cholesterol absorption, cholesterol precursors reflect cholesterol biosynthesis. Here we compare serum and biliary sterol levels in ethnically different populations of patients with gallstone disease (GSD) and stone-free controls to identify differences in cholesterol transport and synthesis between these groups. In this case-control study four cohorts were analyzed: 112 German patients with GSD and 152 controls; two distinct Chilean ethnic groups: Hispanics (100 GSD, 100 controls), and Amerindians (20 GSD, 20 controls); additionally an 8-year follow-up of 70 Hispanics was performed. Serum sterols were measured by gas chromatography / mass spectrometry. Gallbladder bile sterol levels were analyzed in cholesterol GSD and controls. Common ABCG5/8 variants were genotyped. Comparison of serum sterols showed lower levels of phytosterols and higher levels of cholesterol precursors in GSD patients than in controls. The ratios of phytosterols to cholesterol precursors were lower in GSD patients, whereas biliary phytosterol and cholesterol concentrations were elevated as compared with controls. In the follow-up study, serum phytosterol levels were significantly lower even before GSD was detectable by ultrasound. An ethnic gradient in the ratios of phytosterols to cholesterol precursors was apparent (Germans > Hispanics > Amerindians). ABCG5/8 variants did not fully explain the sterol metabolic trait of GSD in any of the cohorts. Individuals predisposed to GSD display increased biliary output of cholesterol in the setting of relatively low intestinal cholesterol absorption, indicating enhanced whole-body sterol clearance. This metabolic trait precedes gallstone formation and is a feature of ethnic groups at higher risk of cholesterol

  1. Intestine-specific MTP and global ACAT2 deficiency lowers acute cholesterol absorption with chylomicrons and HDLs

    Science.gov (United States)

    Boutjdir, Mohamed; Rudel, Lawrence L.; Hussain, M. Mahmood

    2014-01-01

    Intestinal cholesterol absorption involves the chylomicron and HDL pathways and is dependent on microsomal triglyceride transfer protein (MTP) and ABCA1, respectively. Chylomicrons transport free and esterified cholesterol, whereas HDLs transport free cholesterol. ACAT2 esterifies cholesterol for secretion with chylomicrons. We hypothesized that free cholesterol accumulated during ACAT2 deficiency may be secreted with HDLs when chylomicron assembly is blocked. To test this, we studied cholesterol absorption in mice deficient in intestinal MTP, global ACAT2, and both intestinal MTP and global ACAT2. Intestinal MTP ablation significantly increased intestinal triglyceride and cholesterol levels and reduced their transport with chylomicrons. In contrast, global ACAT2 deficiency had no effect on triglyceride absorption but significantly reduced cholesterol absorption with chylomicrons and increased cellular free cholesterol. Their combined deficiency reduced cholesterol secretion with both chylomicrons and HDLs. Thus, contrary to our hypothesis, free cholesterol accumulated in the absence of MTP and ACAT2 is unavailable for secretion with HDLs. Global ACAT2 deficiency causes mild hypertriglyceridemia and reduces hepatosteatosis in mice fed high cholesterol diets by increasing hepatic lipoprotein production by unknown mechanisms. We show that this phenotype is preserved in the absence of intestinal MTP in global ACAT2-deficient mice fed a Western diet. Further, we observed increases in hepatic MTP activity in these mice. Thus, ACAT2 deficiency might increase MTP expression to avoid hepatosteatosis in cholesterol-fed animals. Therefore, ACAT2 inhibition might avert hepatosteatosis associated with high cholesterol diets by increasing hepatic MTP expression and lipoprotein production. PMID:25030663

  2. Intestinal Farnesoid X Receptor Controls Transintestinal Cholesterol Excretion in Mice

    NARCIS (Netherlands)

    de Boer, Jan Freark; Schonewille, Marleen; Boesjes, Marije; Wolters, Henk; Bloks, Vincent W.; Bos, Trijnie; van Dijk, Theo H.; Jurdzinski, Angelika; Boverhof, Renze; Wolters, Justina C.; Kuivenhoven, Jan A.; van Deursen, Jan M.; Oude Elferink, Ronald P. J.; Moschetta, Antonio; Kremoser, Claus; Verkade, Henkjan J.; Kuipers, Folkert; Groen, Albert K.

    2017-01-01

    The role of the intestine in the maintenance of cholesterol homeostasis increasingly is recognized. Fecal excretion of cholesterol is the last step in the atheroprotective reverse cholesterol transport pathway, to which biliary and transintestinal cholesterol excretion (TICE) contribute. The

  3. An overview of membrane transport proteins in Saccharomyces cerevisiae.

    Science.gov (United States)

    Andre, B

    1995-12-01

    All eukaryotic cells contain a wide variety of proteins embedded in the plasma and internal membranes, which ensure transmembrane solute transport. It is now established that a large proportion of these transport proteins can be grouped into families apparently conserved throughout organisms. This article presents the data of an in silicio analysis aimed at establishing a preliminary classification of membrane transport proteins in Saccharomyces cerevisiae. This analysis was conducted at a time when about 65% of all yeast genes were available in public databases. In addition to approximately 60 transport proteins whose function was at least partially known, approximately 100 deduced protein sequences of unknown function display significant sequence similarity to membrane transport proteins characterized in yeast and/or other organisms. While some protein families have been well characterized by classical genetic experimental approaches, others have largely if not totally escaped characterization. The proteins revealed by this in silicio analysis also include a putative K+ channel, proteins similar to aquaporins of plant and animal origin, proteins similar to Na+-solute symporters, a protein very similar to electroneural cation-chloride cotransporters, and a putative Na+-H+ antiporter. A new research area is anticipated: the functional analysis of many transport proteins whose existence was revealed by genome sequencing.

  4. Dietary cholesterol induces trafficking of intestinal Niemann-Pick Type C1 Like 1 from the brush border to endosomes

    DEFF Research Database (Denmark)

    Skov, Marianne; Tønnesen, Carina K; Hansen, Gert H

    2011-01-01

    The transmembrane protein Niemann-Pick C1 Like 1 (NPC1L1) belongs to the Niemann-Pick C1 (NPC1) family of cholesterol transporters and is mainly expressed in the liver and the small intestine. NPC1L1 is believed to be the main transporter responsible for the absorption of dietary cholesterol. Lik...

  5. Bacillus thuringiensis Cyt2Aa2 binding on lipid/cholesterol bilayer depends on protein concentration and time.

    Science.gov (United States)

    Tharad, Sudarat; Moreno-Cencerrado, Alberto; Üzülmez, Öykü; Promdonkoy, Boonhiang; Toca-Herrera, Jose L

    2017-10-14

    Bacillus thuringiensis produces cytolytic proteins (Cyt) that show toxicity against dipteran insect larvae acting directly on the cell membrane. Up to now, two different models have been proposed to explain the interaction mechanism of the cytolytic protein Cyt2Aa2 on lipid membranes: pore-forming and detergent-like action. Here we report on the interaction of Cyt2Aa2 with lipid/cholesterol bilayers at early stage (far from equilibrium) as a function of protein concentration. Quartz crystal microbalance with dissipation (QCM-D) measurements showed that the rate of protein adsorption increased with concentration, although the mass of the final protein-lipid was similar after two hours. In addition, the dissipation (compliance of the hybrid lipid/protein layer) increased with decreasing protein concentration. Furthermore, atomic force microscopy (AFM) revealed that the structure of the protein-lipid layer was concentration and time dependent. A rigid hybrid homogeneous layer was observed at protein concentrations of 50 μg/ml and 100 μg/ml after 30 min. At lower concentrations, 10 μg/ml and 17.5 μg/ml, protein adsorption on the lipid layer led to the formation of small aggregates. Interestingly, at 25 μg/ml a transition of a hole-like structure into a homogeneous layer was observed. This suggests that 25 μg/ml is a threshold concentration for the binding mechanism of Cyt2Aa2 on to lipid membranes. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Effect of growth hormone replacement therapy on plasma lecithin : cholesterol acyltransferase and lipid transfer protein activities in growth hormone-deficient adults

    NARCIS (Netherlands)

    Beentjes, JAM; van Tol, A; Sluiter, WJ; Dullaart, RPF

    The effects of growth hormone (GH) replacement on plasma lecithin:cholesterol acyltransferase (LCAT), cholesteryl ester transfer protein (CETP), and phospholipid transfer protein (PLTP), factors involved in high density lipoprotein (HDL) metabolism, We unknown. We carried out a 6 mouths study in 24

  7. Cholesterol: Its Regulation and Role in Central Nervous System Disorders

    Directory of Open Access Journals (Sweden)

    Matthias Orth

    2012-01-01

    Full Text Available Cholesterol is a major constituent of the human brain, and the brain is the most cholesterol-rich organ. Numerous lipoprotein receptors and apolipoproteins are expressed in the brain. Cholesterol is tightly regulated between the major brain cells and is essential for normal brain development. The metabolism of brain cholesterol differs markedly from that of other tissues. Brain cholesterol is primarily derived by de novo synthesis and the blood brain barrier prevents the uptake of lipoprotein cholesterol from the circulation. Defects in cholesterol metabolism lead to structural and functional central nervous system diseases such as Smith-Lemli-Opitz syndrome, Niemann-Pick type C disease, and Alzheimer’s disease. These diseases affect different metabolic pathways (cholesterol biosynthesis, lipid transport and lipoprotein assembly, apolipoproteins, lipoprotein receptors, and signaling molecules. We review the metabolic pathways of cholesterol in the CNS and its cell-specific and microdomain-specific interaction with other pathways such as the amyloid precursor protein and discuss potential treatment strategies as well as the effects of the widespread use of LDL cholesterol-lowering drugs on brain functions.

  8. Molecular mechanisms of protein-cholesterol interactions in plasma membranes: Functional distinction between topological (tilted) and consensus (CARC/CRAC) domains.

    Science.gov (United States)

    Fantini, Jacques; Di Scala, Coralie; Baier, Carlos J; Barrantes, Francisco J

    2016-09-01

    The molecular mechanisms that control the multiple possible modes of protein association with membrane cholesterol are remarkably convergent. These mechanisms, which include hydrogen bonding, CH-π stacking and dispersion forces, are used by a wide variety of extracellular proteins (e.g. microbial or amyloid) and membrane receptors. Virus fusion peptides penetrate the membrane of host cells with a tilted orientation that is compatible with a transient interaction with cholesterol; this tilted orientation is also characteristic of the process of insertion of amyloid proteins that subsequently form oligomeric pores in the plasma membrane of brain cells. Membrane receptors that are associated with cholesterol generally display linear consensus binding motifs (CARC and CRAC) characterized by a triad of basic (Lys/Arg), aromatic (Tyr/phe) and aliphatic (Leu/Val) amino acid residues. In some cases, the presence of both CARC and CRAC within the same membrane-spanning domain allows the simultaneous binding of two cholesterol molecules, one in each membrane leaflet. In this review the molecular basis and the functional significance of the different modes of protein-cholesterol interactions in plasma membranes are discussed. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Reduction in C-reactive protein and LDL cholesterol and cardiovascular event rates after initiation of rosuvastatin: a prospective study of the JUPITER trial

    DEFF Research Database (Denmark)

    Ridker, Paul M; Danielson, Eleanor; Fonseca, Francisco Ah

    2009-01-01

    BACKGROUND: Statins lower high-sensitivity C-reactive protein (hsCRP) and cholesterol concentrations, and hypothesis generating analyses suggest that clinical outcomes improve in patients given statins who achieve hsCRP concentrations less than 2 mg/L in addition to LDL cholesterol less than 1.......8 mmol/L (LDL cholesterol and hsCRP after the start of statin therapy is controversial. We prospectively tested this hypothesis. METHODS: In an analysis of 15 548 initially healthy men and women participating in the JUPITER trial (87% of full cohort), we...... to on-treatment concentrations of LDL cholesterol (>/=1.8 mmol/L or /=2 mg/L or

  10. Morphology-Variable Aggregates Prepared from Cholesterol-Containing Amphiphilic Glycopolymers: Their Protein Recognition/Adsorption and Drug Delivery Applications

    Directory of Open Access Journals (Sweden)

    Zhao Wang

    2018-02-01

    Full Text Available In this study, a series of diblock glycopolymers, poly(6-O-methacryloyl-d-galactopyranose-b-poly(6-cholesteryloxyhexyl methacrylate (PMAgala-b-PMAChols, with cholesterol/galactose grafts were prepared through a sequential reversible addition-fragmentation chain transfer (RAFT polymerization and deprotection process. The glycopolymers could self-assemble into aggregates with various morphologies depending on cholesterol/galactose-containing block weight ratios, as determined by transmission electronic microscopy (TEM and dynamic laser light scattering (DLS. In addition, the lectin (Ricinus communis agglutinin II, RCA120 recognition and bovine serum albumin (BSA adsorption of the PMAgala-b-PMAChol aggregates were evaluated. The SK-Hep-1 tumor cell inhibition properties of the PMAgala-b-PMAChol/doxorubicin (DOX complex aggregates were further examined in vitro. Results indicate that the PMAgala-b-PMAChol aggregates with various morphologies showed different interaction/recognition features with RCA120 and BSA. Spherical aggregates (d ≈ 92 nm possessed the highest RCA120 recognition ability and lowest BSA protein adsorption. In addition, the DOX-loaded spherical complex aggregates exhibited a better tumor cell inhibition property than those of nanofibrous complex aggregates. The morphology-variable aggregates derived from the amphiphilic glycopolymers may serve as multifunctional biomaterials with biomolecular recognition and drug delivery features.

  11. Elevated levels of preβ1-high-density lipoprotein are associated with cholesterol ester transfer protein, the presence and severity of coronary artery disease.

    Science.gov (United States)

    Bu, Xiao-Min; Niu, Dong-Mei; Wu, Jia; Yuan, Yun-Long; Song, Jia-Xi; Wang, Jun-Jun

    2017-01-10

    Preβ1-high-density lipoprotein (preβ1-HDL), plays an important role in reverse cholesterol transport and exhibits potent risk for coronary artery disease (CAD). However, the association of plasma preβ1-HDL and cholesterol ester transfer protein (CETP) levels in CAD patients and the relationship of preβ1-HDL with extent of CAD are debatable. Preβ1-HDL and CETP levels were measured by enzymed-linked immunosorbent assay (ELISAs) in 88 acute coronary syndromes (ACS), 79 stable coronary artery disease (SCAD) patients and 85 control subjects. The correlation analyses, multiple linear regression analyses and logistic regression analyses were performed, respectively. The preβ1-HDL and CETP levels in ACS patients were significantly higher than those in SCAD patients and both of them were higher than controls'. Preβ1-HDL levels were positively associated with CETP (R = 0.348, P = 0.000), the diameter of stenosis (R = 0.253, P = 0.005), the number of vessel disease (R = 0.274, P = 0.002) and Gensini score (R = 0.227, P = 0.009) in CAD patients. Stepwise multiple linear regression analyses showed that CETP was one of the determinants of preβ1-HDL levels. Logistic regression analysis revealed that elevated preβ1-HDL and CETP were potential risk factors for both ACS and SCAD. The elevated preβ1-HDL levels may change with CETP concentrations in CAD patients and were related to the presence and severity of CAD.

  12. Cholesterol Domains Enhance Transfection

    Science.gov (United States)

    Betker, Jamie L.; Kullberg, Max; Gomez, Joe; Anchordoquy, Thomas J.

    2014-01-01

    The formation of cholesterol domains in lipoplexes has been associated with enhanced serum stability and transfection rates both in cell culture and in vivo. This study utilizes the ability of saturated phosphatidylcholines to promote the formation of cholesterol domains at much lower cholesterol contents than have been utilized in previous work. The results show that lipoplexes with identical cholesterol and cationic lipid contents exhibit significantly improved transfection efficiencies when a domain is present, consistent with previous work. In addition, studies assessing transfection rates in the absence of serum demonstrate that the ability of domains to enhance transfection is not dependent on interactions with serum proteins. Consistent with this hypothesis, characterization of the adsorbed proteins composing the corona of these lipoplex formulations did not reveal a correlation between transfection and the adsorption of a specific protein. Finally, we show that the interaction with serum proteins can promote domain formation in some formulations, and thereby result in enhanced transfection only after serum exposure. PMID:23557286

  13. Hepatitis C Virus Replication Depends on Endosomal Cholesterol Homeostasis.

    Science.gov (United States)

    Stoeck, Ina Karen; Lee, Ji-Young; Tabata, Keisuke; Romero-Brey, Inés; Paul, David; Schult, Philipp; Lohmann, Volker; Kaderali, Lars; Bartenschlager, Ralf

    2018-01-01

    Similar to other positive-strand RNA viruses, hepatitis C virus (HCV) causes massive rearrangements of intracellular membranes, resulting in a membranous web (MW) composed of predominantly double-membrane vesicles (DMVs), the presumed sites of RNA replication. DMVs are enriched for cholesterol, but mechanistic details on the source and recruitment of cholesterol to the viral replication organelle are only partially known. Here we focused on selected lipid transfer proteins implicated in direct lipid transfer at various endoplasmic reticulum (ER)-membrane contact sites. RNA interference (RNAi)-mediated knockdown identified several hitherto unknown HCV dependency factors, such as steroidogenic acute regulatory protein-related lipid transfer domain protein 3 (STARD3), oxysterol-binding protein-related protein 1A and -B (OSBPL1A and -B), and Niemann-Pick-type C1 (NPC1), all residing at late endosome and lysosome membranes and required for efficient HCV RNA replication but not for replication of the closely related dengue virus. Focusing on NPC1, we found that knockdown or pharmacological inhibition caused cholesterol entrapment in lysosomal vesicles concomitant with decreased cholesterol abundance at sites containing the viral replicase factor NS5A. In untreated HCV-infected cells, unesterified cholesterol accumulated at the perinuclear region, partially colocalizing with NS5A at DMVs, arguing for NPC1-mediated endosomal cholesterol transport to the viral replication organelle. Consistent with cholesterol being an important structural component of DMVs, reducing NPC1-dependent endosomal cholesterol transport impaired MW integrity. This suggests that HCV usurps lipid transfer proteins, such as NPC1, at ER-late endosome/lysosome membrane contact sites to recruit cholesterol to the viral replication organelle, where it contributes to MW functionality. IMPORTANCE A key feature of the replication of positive-strand RNA viruses is the rearrangement of the host cell

  14. High protein and cholesterol intakes associated with emergence of glucose intolerance in a low-risk Canadian Inuit population.

    Science.gov (United States)

    Sefidbakht, Saghar; Johnson-Down, Louise; Young, T Kue; Egeland, Grace M

    2016-07-01

    The rate of type 2 diabetes mellitus among Inuit is 12·2 % in individuals over 50 years of age, similar to the Canadian prevalence. Given marked dietary transitions in the Arctic, we evaluated the dietary and other correlates of not previously diagnosed glucose intolerance, defined as type 2 diabetes mellitus, impaired fasting glucose or impaired glucose tolerance. Cross-sectional analyses were limited to adults with a completed 2 h oral glucose tolerance test and without pre-existing diabetes. Anthropometric assessments, health and medication usage questionnaires and a 24 h dietary recall were administered. Canadian International Polar Year Inuit Health Survey (2007-2008). Inuit adults (n 777). Glucose intolerance was associated with older age and adiposity. Percentage of energy from protein above the Acceptable Macronutrient Distribution Range of 35 %, compared with intake within the range, was associated with increased odds of glucose intolerance (OR=1·98; 95 % CI 1·09, 3·61) in multivariable analyses. Further, cholesterol intake in the highest three quartiles combined (median exposures of 207, 416 and 778 mg/d, respectively) compared with the lowest quartile (median intake of 81 mg/d) was associated with glucose intolerance (OR=2·15; 95 % CI 1·23, 3·78) in multivariable analyses. Past-day traditional food consumption was borderline protective of glucose intolerance (P=0·054) and high fibre intake was not significantly protective (P=0·08). The results contribute to the existing literature on high protein and cholesterol intakes as they may relate to diabetes risk.

  15. Cholesterol (image)

    Science.gov (United States)

    Cholesterol is a soft, waxy substance that is present in all parts of the body including the ... and obtained from animal products in the diet. Cholesterol is manufactured in the liver and is needed ...

  16. Cholesterol Test

    Science.gov (United States)

    ... measures: LDL levels. Also known as the "bad" cholesterol, LDL is the main source of blockages in the ... high 240mg/dL and above High LDL (Bad) Cholesterol Level LDL Cholesterol Category Less than 100mg/dL Optimal 100- ...

  17. Inhibition of Cholesterol Synthesis in HepG2 Cells by GINST-Decreasing HMG-CoA Reductase Expression Via AMP-Activated Protein Kinase.

    Science.gov (United States)

    Han, Joon-Seung; Sung, Jong Hwan; Lee, Seung Kwon

    2017-11-01

    GINST, a hydrolyzed ginseng extract, has been reported to have antidiabetic effects and to reduce hyperglycemia and hyperlipidemia. Hypercholesterolemia is caused by diet or genetic factors and can lead to atherosclerosis and coronary heart disease. Thus, the purpose of this study is to determine whether GINST and the ginsenoside metabolite, IH-901 (compound K), reduce cholesterol synthesis in HepG2 cells and the signal transduction pathways involved. Concentrations of cholesterol were measured by using an enzymatic method. Expression levels of sterol regulatory element-binding protein 2 (SREBP2), HMG-CoA reductase (HMGCR), peroxisome proliferators-activated receptor γ (PPARγ), CCAAT/enhancer-binding proteins α (C/EBPα), GAPDH, and phosphorylation of AMP-activated protein kinase α (AMPKα), protein kinase B (PKB, also known as Akt), and mechanistic target of rapamycin complex 1 (mTORC1) were measured using western blot. Total cholesterol concentration decreased after GINST treatment for 24 and 48 h. Expression of HMGCR decreased more with GINST than with the inhibitors, U18666A and atorvastatin, after 48 h in a dose-dependent manner. Phosphorylation of AMPKα increased 2.5x by GINST after 360 min of treatment, and phosphorylation of Akt decreased after 120 and 360 min. We separated compound K from GINST extracts flash chromatography. Compound K decreased cholesterol synthesis in HepG2 cells at 24 and 48 h. Therefore, we conclude that GINST inhibits cholesterol synthesis in HepG2 cells by decreasing HMGCR expression via AMPKα activation. GINST, a hydrolyzed ginseng extract, can inhibit cholesterol synthesis in liver cells via activation of AMPKα. IH-901 (compound K), which is the main component with bioactivity in GINST, also has anticholesterol effects. Thus, we suggest that GINST can be used to reduce hypercholesterolemia. © 2017 Institute of Food Technologists®.

  18. Role of adaptor proteins in motor regulation and membrane transport

    NARCIS (Netherlands)

    M.A. Schlager (Max)

    2010-01-01

    markdownabstract__Abstract__ Active transport along the cytoskeleton is a process essential for proper cellular function. Although much is known about the motor proteins that generate the necessary force and the cytoskeleton that provides the cellular infrastructure, many questions still

  19. Regulatory pathways for ATP-binding cassette transport proteins in kidney proximal tubules

    NARCIS (Netherlands)

    Masereeuw, R.; Russel, F.G.M.

    2012-01-01

    The ATP-binding cassette transport proteins (ABC transporters) represent important determinants of drug excretion. Protective or excretory tissues where these transporters mediate substrate efflux include the kidney proximal tubule. Regulation of the transport proteins in this tissue requires

  20. The effect of cholesteryl ester transfer protein -629C→A promoter polymorphism on high-density lipoprotein cholesterol is dependent on serum triglycerides

    NARCIS (Netherlands)

    S.E. Borggreve (Susanna); H.L. Hillege (Hans); B.H.R. Wolffenbuttel (Bruce); P. de Jong (Paul); S.J.L. Bakker (Stephan); G. van der Steege (Gerrit); A. van Tol (Arie); R.P.F. Dullaart (Robin)

    2005-01-01

    textabstractContext: The -629C→A cholesteryl ester transfer protein (CETP) promoter polymorphism is a determinant of HDL cholesterol (HDL-C). The effect of the closely linked CETP TaqIB polymorphism on HDL-C has been suggested to be modified by obesity and hyperinsulinemia. Objective: Because the

  1. The microsomal triglyceride transfer protein gene-493T variant lowers cholesterol but increases the risk of coronary heart disease.

    Science.gov (United States)

    Ledmyr, Helena; McMahon, Alex D; Ehrenborg, Ewa; Nielsen, Lars B; Neville, Matt; Lithell, Hans; MacFarlane, Peter W; Packard, Christopher J; Karpe, Fredrik

    2004-05-18

    The microsomal triglyceride transfer protein (MTP) transfers lipids into apolipoprotein B-containing lipoproteins for secretion from liver, intestine, and heart. The T-variant of a functional polymorphism in the MTP promoter, MTP-493G/T, has been associated with reduced low-density lipoprotein cholesterol concentrations. We hypothesize that this polymorphism impacts on coronary heart disease (CHD) risk. The effect of the polymorphism was therefore tested in the West of Scotland Coronary Prevention Study biobank (580 cases and 1160 controls). MTP-493T carrier status was associated with significantly increased risk of CHD despite a small reduction in total cholesterol. Compared with the genotypic group with the lowest event rate (MTP-493GG, pravastatin treatment), the respective odds ratios (95% confidence interval) in the placebo group for CHD events were: GG, 1.23 (0.92 to 1.63); GT, 1.53 (1.12 to 2.08); and TT, 2.78 (1.53 to 5.05), suggestive of a gene-dose effect. The excess risk for CHD of the MTP-493T-variant was eliminated by pravastatin treatment. The Uppsala Longitudinal Study of Adult Men (ULSAM), which is a 20-year follow-up study of CHD, was used as an independent confirmatory database. These unexpected findings prompted the investigation of non-plasma lipid factors that could associate the MTP gene with CHD risk. In a limited number of subjects (n=18), heart muscle biopsies showed a MTP-493T genotype-specific depression of MTP mRNA expression. The MTP-493T variant confers an increased risk of CHD that is unrelated to plasma lipids and lipoproteins, but eliminated by pravastatin treatment. A direct effect of the MTP polymorphism on myocardial lipid metabolism and vulnerability upon ischemic damage cannot be excluded.

  2. Protein transport across the small intestine in food allergy.

    Science.gov (United States)

    Reitsma, Marit; Westerhout, Joost; Wichers, Harry J; Wortelboer, Heleen M; Verhoeckx, Kitty C M

    2014-01-01

    In view of the imminent deficiency of protein sources for human consumption in the near future, new protein sources need to be identified. However, safety issues such as the risk of allergenicity are often a bottleneck, due to the absence of predictive, validated and accepted methods for risk assessment. The current strategy to assess the allergenic potential of proteins focuses mainly on homology, stability and cross-reactivity, although other factors such as intestinal transport might be of added value too. In this review, we present an overview of the knowledge of protein transport across the intestinal wall and the methods currently being used to measure this. A literature study reveals that protein transport in sensitised persons occurs para-cellularly with the involvement of mast cells, and trans-cellularly via enterocytes, while in non-sensitised persons micro-fold cells and enterocytes are considered most important. However, there is a lack of comparable systematic studies on transport of allergenic proteins. Knowledge of the multiple protein transport pathways and which model system can be useful to study these processes may be of added value in the risk assessment of food allergenicity. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Biomimetic materials for protein storage and transport

    Science.gov (United States)

    Firestone, Millicent A [Elmhurst, IL; Laible, Philip D [Villa Park, IL

    2012-05-01

    The invention provides a method for the insertion of protein in storage vehicles and the recovery of the proteins from the vehicles, the method comprising supplying isolated protein; mixing the isolated protein with a fluid so as to form a mixture, the fluid comprising saturated phospholipids, lipopolymers, and a surfactant; cycling the mixture between a first temperature and a second temperature; maintaining the mixture as a solid for an indefinite period of time; diluting the mixture in detergent buffer so as to disrupt the composition of the mixture, and diluting to disrupt the fluid in its low viscosity state for removal of the guest molecules by, for example, dialysis, filtering or chromatography dialyzing/filtering the emulsified solid.

  4. Mucins and calcium phosphate precipitates additively stimulate cholesterol crystallization

    NARCIS (Netherlands)

    van den Berg, A. A.; van Buul, J. D.; Tytgat, G. N.; Groen, A. K.; Ostrow, J. D.

    1998-01-01

    Human biliary mucin and calcium binding protein (CBP) influence formation of both calcium salt precipitates and cholesterol crystals and colocalize in the center of cholesterol gallstones. We investigated how physiological concentrations of these proteins regulate cholesterol crystallization in

  5. Placenta Copper Transport Proteins in Preeclampsia

    Science.gov (United States)

    Placental insufficiency underlying preeclampsia (PE) is associated with impaired placental angiogenesis. As copper (Cu) is essential to angiogenesis, we investigated differences in the expression of placental Cu transporters Menkes (ATP7A), Wilsons (ATP7B) and the Cu chaperone (CCS) for superoxide d...

  6. CER-001, a HDL-mimetic, stimulates the reverse lipid transport and atherosclerosis regression in high cholesterol diet-fed LDL-receptor deficient mice.

    Science.gov (United States)

    Tardy, Claudine; Goffinet, Marine; Boubekeur, Nadia; Ackermann, Rose; Sy, Gavin; Bluteau, Alice; Cholez, Guy; Keyserling, Constance; Lalwani, Narendra; Paolini, John F; Dasseux, Jean-Louis; Barbaras, Ronald; Baron, Rudi

    2014-01-01

    CER-001 is a novel engineered HDL-mimetic comprised of recombinant human apoA-I and phospholipids that was designed to mimic the beneficial properties of nascent pre-β HDL. In this study, we have evaluated the capacity of CER-001 to perform reverse lipid transport in single dose studies as well as to regress atherosclerosis in LDLr(-/-) mice after short-term multiple-dose infusions. CER-001 induced cholesterol efflux from macrophages and exhibited anti-inflammatory response similar to natural HDL. Studies with HUVEC demonstrated CER-001 at a concentration of 500 μg/mL completely suppressed the secretion of cytokines IL-6, IL-8, GM-CSF and MCP-1. Following infusion of CER-001 (10mg/kg) in C57Bl/6J mice, we observed a transient increase in the mobilization of unesterified cholesterol in HDL particles containing recombinant human apoA-I. Finally we show that cholesterol elimination was stimulated in CER-001 treated animals as demonstrated by the increased cholesterol concentration in liver and feces. In a familial hypercholesterolemia mouse model (LDL-receptor deficient mice), the infusion of CER-001 caused 17% and 32% reductions in plaque size, 17% and 23% reductions in lipid content after 5 and 10 doses given every 2 days, respectively. Also, there was an 80% reduction in macrophage content in the plaque following 5 doses, and decreased VCAM-1 expression by 16% and 22% in the plaque following 5 and 10 intravenous doses of CER-001, respectively. These data demonstrate that CER-001 rapidly enhances reverse lipid transport in the mouse, reducing vascular inflammation and promoting regression of diet-induced atherosclerosis in LDLr(-/-) mice upon a short-term multiple dose treatment. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  7. Mechanistic logic underlying the axonal transport of cytosolic proteins

    Science.gov (United States)

    Scott, David A.; Das, Utpal; Tang, Yong; Roy, Subhojit

    2011-01-01

    Proteins vital to presynaptic function are synthesized in the neuronal perikarya and delivered into synapses via two modes of axonal transport. While membrane-anchoring proteins are conveyed in fast axonal transport via motor-driven vesicles, cytosolic proteins travel in slow axonal transport; via mechanisms that are poorly understood. We found that in cultured axons, populations of cytosolic proteins tagged to photoactivable-GFP (PA-GFP) move with a slow motor-dependent anterograde bias; distinct from vesicular-trafficking or diffusion of untagged PA-GFP. The overall bias is likely generated by an intricate particle-kinetics involving transient assembly and short-range vectorial spurts. In-vivo biochemical studies reveal that cytosolic proteins are organized into higher-order structures within axon-enriched fractions that are largely segregated from vesicles. Data-driven biophysical modeling best predicts a scenario where soluble molecules dynamically assemble into mobile supra-molecular structures. We propose a model where cytosolic proteins are transported by dynamically assembling into multi-protein complexes that are directly/indirectly conveyed by motors. PMID:21555071

  8. The PIN-FORMED (PIN) protein family of auxin transporters.

    Science.gov (United States)

    Krecek, Pavel; Skupa, Petr; Libus, Jirí; Naramoto, Satoshi; Tejos, Ricardo; Friml, Jirí; Zazímalová, Eva

    2009-01-01

    The PIN-FORMED (PIN) proteins are secondary transporters acting in the efflux of the plant signal molecule auxin from cells. They are asymmetrically localized within cells and their polarity determines the directionality of intercellular auxin flow. PIN genes are found exclusively in the genomes of multicellular plants and play an important role in regulating asymmetric auxin distribution in multiple developmental processes, including embryogenesis, organogenesis, tissue differentiation and tropic responses. All PIN proteins have a similar structure with amino- and carboxy-terminal hydrophobic, membrane-spanning domains separated by a central hydrophilic domain. The structure of the hydrophobic domains is well conserved. The hydrophilic domain is more divergent and it determines eight groups within the protein family. The activity of PIN proteins is regulated at multiple levels, including transcription, protein stability, subcellular localization and transport activity. Different endogenous and environmental signals can modulate PIN activity and thus modulate auxin-distribution-dependent development. A large group of PIN proteins, including the most ancient members known from mosses, localize to the endoplasmic reticulum and they regulate the subcellular compartmentalization of auxin and thus auxin metabolism. Further work is needed to establish the physiological importance of this unexpected mode of auxin homeostasis regulation. Furthermore, the evolution of PIN-based transport, PIN protein structure and more detailed biochemical characterization of the transport function are important topics for further studies.

  9. Immune regulation of Rab proteins expression and intracellular transport.

    Science.gov (United States)

    Pei, Gang; Bronietzki, Marc; Gutierrez, Maximiliano Gabriel

    2012-07-01

    Compartmentalization in cells of the immune system, the focus of this review, facilitates the spatiotemporal organization of cellular responses essential for specialized immune functions. In this process of compartment maintenance, Rab proteins are central regulators of protein-mediated transport and fusion of intracellular structures. It is widely believed that the intracellular concentration of proteins that regulate intracellular transport, including Rab proteins, is constitutively mantained. However, there is a growing body of evidence indicating that transcriptional rates of Rab proteins can be modified. This process is especially evident during immune activation and argues that after activation, these cells require higher levels of Rab proteins. The aim of this review is to discuss evidence showing the increasing links between Rab protein expression and intracellular transport, particularly in monocytes and macrophages. We highlight here biological processes in which the expression of Rab GTPases is selectively regulated, leading to the activation of specific intracellular routes. Further, we focus on the immune regulation of intracellular transport after cytokine activation and microbial infection, with an emphasis in mycobacterial infection.

  10. Dietary fiber prevents obesity-related liver lipotoxicity by modulating sterol-regulatory element binding protein pathway in C57BL/6J mice fed a high-fat/cholesterol diet.

    Science.gov (United States)

    Han, Shufen; Jiao, Jun; Zhang, Wei; Xu, Jiaying; Wan, Zhongxiao; Zhang, Weiguo; Gao, Xiaoran; Qin, Liqiang

    2015-10-29

    Adequate intake of dietary fibers has proven metabolic and cardiovascular benefits, molecular mechanisms remain still limited. This study was aimed to investigate the effects of cereal dietary fiber on obesity-related liver lipotoxicity in C57BL/6J mice fed a high-fat/cholesterol (HFC) diet and underlying mechanism. Forty-eight adult male C57BL/6J mice were randomly given a reference chow diet, or a high fat/cholesterol (HFC) diet supplemented with or without oat fiber or wheat bran fiber for 24 weeks. Our results showed mice fed oat or wheat bran fiber exhibited lower weight gain, lipid profiles and insulin resistance, compared with HFC diet. The two cereal dietary fibers potently decreased protein expressions of sterol regulatory element binding protein-1 and key factors involved in lipogenesis, including fatty acid synthase and acetyl-CoA carboxylase in target tissues. At molecular level, the two cereal dietary fibers augmented protein expressions of peroxisome proliferator-activated receptor alpha and gamma, liver X receptor alpha, and ATP-binding cassette transporter A1 in target tissues. Our findings indicated that cereal dietary fiber supplementation abrogated obesity-related liver lipotoxicity and dyslipidemia in C57BL/6J mice fed a HFC diet. In addition, the efficacy of oat fiber is greater than wheat bran fiber in normalizing these metabolic disorders and pathological profiles.

  11. STARD4 knockdown in HepG2 cells disrupts cholesterol trafficking associated with the plasma membrane, ER, and ERC

    DEFF Research Database (Denmark)

    Garbarino, J.; Pan, M. H.; Chin, H. F.

    2012-01-01

    STARD4, a member of the evolutionarily conserved START gene family, has been implicated in the non-vesicular intracellular transport of cholesterol. However, the direction of transport and the membranes with which this protein interacts are not clear. We present studies of STARD4 function using...... small hairpin RNA knockdown technology to reduce STARD4 expression in HepG2 cells. In a cholesterol-poor environment, we found that a reduction in STARD4 expression leads to retention of cholesterol at the plasma membrane, reduction of endoplasmic reticulum-associated cholesterol, and decreased ACAT...... synthesized cholesteryl esters. Furthermore, D4 KD cells exhibited a reduced rate of sterol transport to the endocytic recycling compartment after cholesterol repletion. Although these cells displayed normal endocytic trafficking in cholesterol-poor and replete conditions, cell surface low density lipoprotein...

  12. DIETARY FISH OIL-INDUCED CHANGES IN INTRAHEPATIC CHOLESTEROL TRANSPORT AND BILE-ACID SYNTHESIS IN RATS

    NARCIS (Netherlands)

    SMIT, MJ; TEMMERMAN, AM; WOLTERS, H; KUIPERS, F; BEYNEN, AC; VONK, RJ

    Hepatic cholesterol metabolism was studied in rats fed purified diets supplemented (9% wt/wt) with either fish oil (FO) (n-3 fatty acids) or corn oil (CO) (n-6 fatty acids) for 4 wk. Rats were equipped with permanent catheters in heart, bile duct, and duodenum to allow studies under normal feeding

  13. Association among retinol-binding protein 4, small dense LDL cholesterol and oxidized LDL levels in dyslipidemia subjects.

    Science.gov (United States)

    Wu, Jia; Shi, Yong-hui; Niu, Dong-mei; Li, Han-qing; Zhang, Chun-ni; Wang, Jun-jun

    2012-06-01

    To investigate retinol-binding protein 4 (RBP4), small dense low-density lipoprotein cholesterol (sdLDL-C) and oxidized low-density lipoprotein (ox-LDL) levels and their associations in dyslipidemia subjects. We determined RBP4, sdLDL-C, ox-LDL levels in 150 various dyslipidemia subjects and 50 controls. The correlation analysis and multiple linear regression analysis were performed. The RBP4, sdLDL-C and ox-LDL levels were found increased in various dyslipidemia subjects. The sdLDL-C levels were positively correlated with RBP4 (r=0.273, P=0.001) and ox-LDL (r=0.273, P=0.001). RBP4 levels were also correlated with ox-LDL (r=0.167, P=0.043). The multiple regression analysis showed that only sdLDL-C was a significant independent predictor for RBP4 (β coefficient=0.219, P=0.009; adjusted R(2)=0.041) and ox-LDL (β coefficient=0.253, P=0.003; adjusted R(2)=0.057) levels, respectively. The independent associations of sdLDL-C with RBP4 and ox-LDL were observed in dyslipidemia subjects. RBP4 may play an important role in lipid metabolism of atherosclerosis, particularly in formation of sdLDL. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  14. Cholesterol absorption inhibitors as a therapeutic option for hypercholesterolaemia.

    Science.gov (United States)

    Burnett, John R; Huff, Murray W

    2006-11-01

    The development of cholesterol-lowering drugs (including a variety of statins, bile acid-binding resins and recently discovered inhibitors of cholesterol absorption) has expanded the options for cardiovascular prevention. Recent treatment guidelines emphasise that individuals at substantial risk for atherosclerotic coronary heart disease should meet defined targets for LDL cholesterol concentrations. Combination therapy with drugs that have different or complementary mechanisms of action is often needed to achieve lipid goals. Existing approaches to the treatment of hypercholesterolaemia are still ineffective in halting the progression of coronary artery disease in some patients despite combination therapies. Other patients are resistant to conventional drug treatment and remain at high risk for the development and progression of atherosclerotic cardiovascular disease and alternative approaches are needed. The discovery and development of ezetimibe (a novel, selective and potent cholesterol absorption inhibitor) has advanced the treatment of hypercholesterolaemia. New agents including the phytostanol preparation FM-VP4 and inhibitors of acyl coenzyme A:cholesterol acyltransferase, the apical Na(+)-dependent bile acid transporter and microsomal triglyceride transfer protein may also play a future role in combination therapy. This review focuses on the recent progress in the molecular mechanisms of intestinal cholesterol absorption and transport, and novel therapeutic approaches to inhibit the cholesterol absorption process.

  15. Cholesterol regulates contractility and inotropic response to β2-adrenoceptor agonist in the mouse atria: Involvement of Gi-protein-Akt-NO-pathway.

    Science.gov (United States)

    Odnoshivkina, Yulia G; Sytchev, Vaycheslav I; Petrov, Alexey M

    2017-06-01

    Majority of cardiac β2-adrenoceptors is located in cholesterol-rich microdomains. Here, we have investigated the underlying mechanisms by which a slight to moderate cholesterol depletion with methyl-β-cyclodextrin (MβCD, 1 and 5mM) interferes with contractility and inotropic effect of β2-adrenergic agonist (fenoterol, 50μM) in the mouse atria. Treatment with MβCD itself increased amplitude of Ca 2+ transient but did not change the contraction amplitude due to a clamping action of elevated NO. Cholesterol depletion significantly attenuated the positive inotropic response to fenoterol which is accompanied by increase in NO generation and decrease in Ca 2+ transient. Influence of 1mM MβCD on the fenoterol-driven changes in both contractility and NO level was strongly attenuated by inhibition of G i -protein (pertussis toxin), Akt (Akt 1/2 kinase inhibitor) or NO-synthase (L-NAME). After exposure to 5mM MβCD, pertussis toxin or Akt inhibitor could recover the β2-agonist effects on contractility, NO production and Ca 2+ transient, while L-NAME only reduced NO level. An adenylyl cyclase activator (forskolin, 50nM) had no influence on the MβCD-induced changes in the β2-agonist effects. Obtained results suggest that slight cholesterol depletion upregulates G i -protein/Akt/NO-synthase signaling that attenuates the positive inotropic response to β2-adrenergic stimulation without altering the Ca 2+ transient. Whilst moderate cholesterol depletion additionally could suppress the enhancement of the Ca 2+ transient amplitude caused by the β2-adrenergic agonist administration in G i -protein/Akt-dependent but NO-independent manner. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Microvillus-Specific Protein Tyrosine Phosphatase SAP-1 Plays a Role in Regulating the Intestinal Paracellular Transport of Macromolecules.

    Science.gov (United States)

    Mori, Shingo; Kamei, Noriyasu; Murata, Yoji; Takayama, Kozo; Matozaki, Takashi; Takeda-Morishita, Mariko

    2017-09-01

    The stomach cancer-associated protein tyrosine phosphatase 1 (SAP-1) is a receptor-type protein tyrosine phosphatase that is specifically expressed on the apical membrane of the intestinal epithelium. SAP-1 is known to maintain the balance of phosphorylation of proteins together with protein kinases; however, its biological function and impact on pharmacokinetics in the intestine remain unclear. The present study, therefore, aimed at clarifying the relationship between SAP-1 and the intestinal absorption behaviors of typical transporter substrates and macromolecules. The endogenous levels of glucose and total cholesterol in the blood were similar between wild-type and SAP-1-deficient mice (Sap1 -/- ), suggesting no contribution of SAP-1 to biogenic influx. Moreover, in vitro transport study with everted ileal sacs demonstrated that there was no difference in the absorption of breast cancer resistance protein, P-glycoprotein, and peptide transporter substrates between both mice. However, absorptive clearance of macromolecular model dextrans (FD-4 and FD-10) in Sap1 -/- mice was significantly higher than that in wild-type mice, and this was confirmed by the trend of increased FD-4 absorption from colonic loops of Sap1 -/- mice. Therefore, the results of this study suggest the partial contribution of SAP-1 to the regulated transport of hydrophilic macromolecules through paracellular tight junctions. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  17. Intracellular transport proteins: classification, structure and function of kinesins

    Directory of Open Access Journals (Sweden)

    Agnieszka Chudy

    2011-09-01

    Full Text Available Correct cell functioning, division and morphogenesis rely on efficient intracellular transport. Apart from dyneins and myosins, kinesins are the main proteins responsible for intracellular movement. Kinesins are a large, diverse group of motor proteins, which based on phylogenetic similarity were classified into fourteen families. Among these families, due to the location of their motor domains, three groups have been characterized: N-, C- and M-kinesin. As molecular motors, kinesins transport various molecules and vesicles mainly towards the microtubule plus end (from the cell body participating in anterograde transport, although there are also kinesins involved in retrograde transport (C-kinesins. Kinesins are also involved in spindle formation, chromosome segregation, and spermatogenesis. Because of their great importance for the correct functioning of cells, mutations in kinesin coding genes may lead to such neurodegenerative diseases as dominant hereditary spastic paraplegia or Charcot-Marie-Tooth disease.

  18. Position-dependent Effects of Polylysine on Sec Protein Transport*

    Science.gov (United States)

    Liang, Fu-Cheng; Bageshwar, Umesh K.; Musser, Siegfried M.

    2012-01-01

    The bacterial Sec protein translocation system catalyzes the transport of unfolded precursor proteins across the cytoplasmic membrane. Using a recently developed real time fluorescence-based transport assay, the effects of the number and distribution of positive charges on the transport time and transport efficiency of proOmpA were examined. As expected, an increase in the number of lysine residues generally increased transport time and decreased transport efficiency. However, the observed effects were highly dependent on the polylysine position in the mature domain. In addition, a string of consecutive positive charges generally had a more significant effect on transport time and efficiency than separating the charges into two or more charged segments. Thirty positive charges distributed throughout the mature domain resulted in effects similar to 10 consecutive charges near the N terminus of the mature domain. These data support a model in which the local effects of positive charge on the translocation kinetics dominate over total thermodynamic constraints. The rapid translocation kinetics of some highly charged proOmpA mutants suggest that the charge is partially shielded from the electric field gradient during transport, possibly by the co-migration of counter ions. The transport times of precursors with multiple positively charged sequences, or “pause sites,” were fairly well predicted by a local effect model. However, the kinetic profile predicted by this local effect model was not observed. Instead, the transport kinetics observed for precursors with multiple polylysine segments support a model in which translocation through the SecYEG pore is not the rate-limiting step of transport. PMID:22367204

  19. High blood cholesterol levels

    Science.gov (United States)

    Cholesterol - high; Lipid disorders; Hyperlipoproteinemia; Hyperlipidemia; Dyslipidemia; Hypercholesterolemia ... There are many types of cholesterol. The ones talked about most are: ... lipoprotein (HDL) cholesterol -- often called "good" cholesterol ...

  20. The Ala54Thr Polymorphism of the Fatty Acid Binding Protein 2 Gene Modulates HDL Cholesterol in Mexican-Americans with Type 2 Diabetes

    Directory of Open Access Journals (Sweden)

    Lorena M. Salto

    2015-12-01

    Full Text Available The alanine to threonine amino acid substitution at codon 54 (Ala54Thr of the intestinal fatty acid binding protein (FABP2 has been associated with elevated levels of insulin and blood glucose as well as with dyslipidemia. The aim of this study was to characterize the effect of this FABP2 polymorphism in Mexican-Americans with type 2 diabetes (T2D in the context of a three-month intervention to determine if the polymorphism differentially modulates selected clinical outcomes. For this study, we genotyped 43 participant samples and performed post-hoc outcome analysis of the profile changes in fasting blood glucose, HbA1c, insulin, lipid panel and body composition, stratified by the Ala54Thr polymorphism. Our results show that the Thr54 allele carriers (those who were heterozygous or homozygous for the threonine-encoding allele had lower HDL cholesterol and higher triglyceride levels at baseline compared to the Ala54 homozygotes (those who were homozygous for the alanine-encoding allele. Both groups made clinically important improvements in lipid profiles and glycemic control as a response to the intervention. Whereas the Ala54 homozygotes decreased HDL cholesterol in the context of an overall total cholesterol decrease, Thr54 allele carriers increased HDL cholesterol as part of an overall total cholesterol decrease. We conclude that the Ala54Thr polymorphism of FABP2 modulates HDL cholesterol in Mexican-Americans with T2D and that Thr54 allele carriers may be responsive in interventions that include dietary changes.

  1. Ezetimibe Increases Endogenous Cholesterol Excretion in Humans.

    Science.gov (United States)

    Lin, Xiaobo; Racette, Susan B; Ma, Lina; Wallendorf, Michael; Ostlund, Richard E

    2017-05-01

    Ezetimibe improves cardiovascular outcomes when added to optimum statin treatment. It lowers low-density lipoprotein cholesterol and percent intestinal cholesterol absorption, but the exact cardioprotective mechanism is unknown. We tested the hypothesis that the dominant effect of ezetimibe is to increase the reverse transport of cholesterol from rapidly mixing endogenous cholesterol pool into the stool. In a randomized, placebo-controlled, double-blind parallel trial in 24 healthy subjects with low-density lipoprotein cholesterol 100 to 200 mg/dL, we measured cholesterol metabolism before and after a 6-week treatment period with ezetimibe 10 mg/d or placebo. Plasma cholesterol was labeled by intravenous infusion of cholesterol-d 7 in a lipid emulsion and dietary cholesterol with cholesterol-d 5 and sitostanol-d 4 solubilized in oil. Plasma and stool samples collected during a cholesterol- and phytosterol-controlled metabolic kitchen diet were analyzed by mass spectrometry. Ezetimibe reduced intestinal cholesterol absorption efficiency 30±4.3% (SE, P <0.0001) and low-density lipoprotein cholesterol 19.8±1.9% ( P =0.0001). Body cholesterol pool size was unchanged, but fecal endogenous cholesterol excretion increased 66.6±12.2% ( P <0.0001) and percent cholesterol excretion from body pools into the stool increased 74.7±14.3% ( P <0.0001), whereas plasma cholesterol turnover rose 26.2±3.6% ( P =0.0096). Fecal bile acids were unchanged. Ezetimibe increased the efficiency of reverse cholesterol transport from rapidly mixing plasma and tissue pools into the stool. Further work is needed to examine the potential relation of reverse cholesterol transport and whole body cholesterol metabolism to coronary events and the treatment of atherosclerosis. URL: http://www.clinicaltrials.gov. Unique identifier: NCT01603758. © 2017 American Heart Association, Inc.

  2. Ion transport across the biological membrane by computational protein design

    Science.gov (United States)

    Grigoryan, Gevorg

    The cellular membrane is impermeable to most of the chemicals the cell needs to take in or discard to survive. Therefore, transporters-a class of transmembrane proteins tasked with shuttling cargo chemicals in and out of the cell-are essential to all cellular life. From existing crystal structures, we know transporters to be complex machines, exquisitely tuned for specificity and controllability. But how could membrane-bound life have evolved if it needed such complex machines to exist first? To shed light onto this question, we considered the task of designing a transporter de novo. As our guiding principle, we took the ``alternating-access model''-a conceptual mechanism stating that transporters work by rocking between two conformations, each exposing the cargo-binding site to either the intra- or the extra-cellular environment. A computational design framework was developed to encode an anti-parallel four-helix bundle that rocked between two alternative states to orchestrate the movement of Zn(II) ions across the membrane. The ensemble nature of both states was accounted for using a free energy-based approach, and sequences were chosen based on predicted formation of the targeted topology in the membrane and bi-stability. A single sequence was prepared experimentally and shown to function as a Zn(II) transporter in lipid vesicles. Further, transport was specific to Zn(II) ions and several control peptides supported the underlying design principles. This included a mutant designed to retain all properties but with reduced rocking, which showed greatly depressed transport ability. These results suggest that early transporters could have evolved in the context of simple topologies, to be later tuned by evolution for improved properties and controllability. Our study also serves as an important advance in computational protein design, showing the feasibility of designing functional membrane proteins and of tuning conformational landscapes for desired function

  3. Protein replacement therapy partially corrects the cholesterol-storage phenotype in a mouse model of Niemann-Pick type C2 disease.

    Directory of Open Access Journals (Sweden)

    Gitte Krogh Nielsen

    Full Text Available Niemann-Pick type C2 (NPC2 disease is a fatal autosomal recessive neurovisceral degenerative disorder characterized by late endosomal-lysosomal sequestration of low-density lipoprotein derived cholesterol. The breach in intracellular cholesterol homeostasis is caused by deficiency of functional NPC2, a soluble sterol binding protein targeted to the lysosomes by binding the mannose-6-phosphate receptor. As currently there is no effective treatment for the disorder, we have investigated the efficacy of NPC2 replacement therapy in a murine gene-trap model of NPC2-disease generated on the 129P2/OlaHsd genetic background. NPC2 was purified from bovine milk and its functional competence assured in NPC2-deficient fibroblasts using the specific cholesterol fluorescent probe filipin. For evaluation of phenotype correction in vivo, three-week-old NPC2(-/- mice received two weekly intravenous injections of 5 mg/kg NPC2 until trial termination 66 days later. Whereas the saline treated NPC2(-/- mice exhibited massive visceral cholesterol storage as compared to their wild-type littermates, administration of NPC2 caused a marked reduction in cholesterol build up. The histological findings, indicating an amelioration of the disease pathology in liver, spleen, and lungs, corroborated the biochemical results. Little or no difference in the overall cholesterol levels was observed in the kidneys, blood, cerebral cortex and hippocampus when comparing NPC2(-/- and wild type mice. However, cerebellum cholesterol was increased about two fold in NPC2(-/- mice compared with wild-type littermates. Weight gain performance was slightly improved as a result of the NPC2 treatment but significant motor coordination deficits were still observed. Accordingly, ultrastructural cerebellar abnormalities were detected in both saline treated and NPC2 treated NPC2(-/- animals 87 days post partum. Our data indicate that protein replacement may be a beneficial therapeutic approach in the

  4. Rab proteins: The key regulators of intracellular vesicle transport

    International Nuclear Information System (INIS)

    Bhuin, Tanmay; Roy, Jagat Kumar

    2014-01-01

    Vesicular/membrane trafficking essentially regulates the compartmentalization and abundance of proteins within the cells and contributes in many signalling pathways. This membrane transport in eukaryotic cells is a complex process regulated by a large and diverse array of proteins. A large group of monomeric small GTPases; the Rabs are essential components of this membrane trafficking route. Most of the Rabs are ubiquitously expressed proteins and have been implicated in vesicle formation, vesicle motility/delivery along cytoskeleton elements and docking/fusion at target membranes through the recruitment of effectors. Functional impairments of Rabs affecting transport pathways manifest different diseases. Rab functions are accompanied by cyclical activation and inactivation of GTP-bound and GDP-bound forms between the cytosol and membranes which is regulated by upstream regulators. Rab proteins are characterized by their distinct sub-cellular localization and regulate a wide variety of endocytic, transcytic and exocytic transport pathways. Mutations of Rabs affect cell growth, motility and other biological processes. - Highlights: • Rab proteins regulate different signalling pathways. • Deregulation of Rabs is the fundamental causes of a variety of human diseases. • This paper gives potential directions in developing therapeutic targets. • This paper also gives ample directions for modulating pathways central to normal physiology. • These are the huge challenges for drug discovery and delivery in near future

  5. Rab proteins: The key regulators of intracellular vesicle transport

    Energy Technology Data Exchange (ETDEWEB)

    Bhuin, Tanmay [Cell and Developmental Biology Unit, Department of Zoology, The University of Burdwan, Golapbag 713104 (India); Roy, Jagat Kumar, E-mail: jkroy@bhu.ac.in [Cytogenetics Laboratory, Department of Zoology, Banaras Hindu University, Varanasi 221005 (India)

    2014-10-15

    Vesicular/membrane trafficking essentially regulates the compartmentalization and abundance of proteins within the cells and contributes in many signalling pathways. This membrane transport in eukaryotic cells is a complex process regulated by a large and diverse array of proteins. A large group of monomeric small GTPases; the Rabs are essential components of this membrane trafficking route. Most of the Rabs are ubiquitously expressed proteins and have been implicated in vesicle formation, vesicle motility/delivery along cytoskeleton elements and docking/fusion at target membranes through the recruitment of effectors. Functional impairments of Rabs affecting transport pathways manifest different diseases. Rab functions are accompanied by cyclical activation and inactivation of GTP-bound and GDP-bound forms between the cytosol and membranes which is regulated by upstream regulators. Rab proteins are characterized by their distinct sub-cellular localization and regulate a wide variety of endocytic, transcytic and exocytic transport pathways. Mutations of Rabs affect cell growth, motility and other biological processes. - Highlights: • Rab proteins regulate different signalling pathways. • Deregulation of Rabs is the fundamental causes of a variety of human diseases. • This paper gives potential directions in developing therapeutic targets. • This paper also gives ample directions for modulating pathways central to normal physiology. • These are the huge challenges for drug discovery and delivery in near future.

  6. Study of the transport of mercurial compounds by seric proteins

    International Nuclear Information System (INIS)

    Jullien-Saint Guily, Nicole

    1970-01-01

    A bond between the seric proteins and various mercurial compounds labeled with the radioisotopes 203 Hg and 197 Hg was demonstrated by means of research methods specific to radioactivity combined with protein separation techniques. In the course of this study it was shown how strongly the composition of the buffer during electrophoretic migration influences the transport of certain organo-mercurial compounds by the seric proteins. By means of a thioloprive: N - ethyl - maleimide, labeled with 14 C, it was proved that the bonding sites between the proteins and the mercurial compounds were the thiol groups of the proteins but that other bonding sites, in particular the amino groups, could also be involved. (author) [fr

  7. Relationship between circulating microRNA-30c with total- and LDL-cholesterol, their circulatory transportation and effect of statins.

    Science.gov (United States)

    Sodi, Ravinder; Eastwood, Jarlath; Caslake, Muriel; Packard, Chris J; Denby, Laura

    2017-03-01

    Small non-coding microRNAs (miR) have important regulatory roles and are used as biomarkers of disease. We investigated the relationship between lipoproteins and circulating miR-30c, evaluated how they are transported in circulation and determined whether statins altered the circulating concentration of miR-30c. To determine the relationship between lipoproteins and circulating miR-30c, serum samples from 79 subjects recruited from a lipid clinic were evaluated. Ultracentrifugation and nanoparticle tracking analysis was used to evaluate the transportation of miR-30c in the circulation by lipoproteins and extracellular vesicles in three healthy volunteers. Using archived samples from previous studies, the effects of 40mg rosuvastatin (n=22) and 40mg pravastatin (n=24) on miR-30c expression was also examined. RNA extraction, reverse transcription-quantitative real-time polymerase chain reaction was carried out using standard procedures. When stratified according to total cholesterol concentration, there was increased miR-30c expression in the highest compared to the lowest tertile (p=0.035). There was significant positive correlation between miR-30c and total- (r=0.367; p=0.002) and LDL-cholesterol (r=0.391; p=0.001). We found that miR-30c was transported in both exosomes and on HDL3. There was a 3.8-fold increased expression of circulating miR-30c after pravastatin treatment for 1year (p=0.005) but no significant change with atorvastatin after 8weeks (p=0.145). This study shows for the first-time in humans that circulating miR-30c is significantly, positively correlated with total- and LDL-cholesterol implicating regulatory functions in lipid homeostasis. We show miR-30c is transported in both exosomes and on HDL3 and pravastatin therapy significantly increased circulating miR-30c expression adding to the pleiotropic dimensions of statins. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Deletion of Macrophage Vitamin D Receptor Promotes Insulin Resistance and Monocyte Cholesterol Transport to Accelerate Atherosclerosis in Mice

    Directory of Open Access Journals (Sweden)

    Jisu Oh

    2015-03-01

    Full Text Available Intense effort has been devoted to understanding predisposition to chronic systemic inflammation because it contributes to cardiometabolic disease. We demonstrate that deletion of the macrophage vitamin D receptor (VDR in mice (KODMAC is sufficient to induce insulin resistance by promoting M2 macrophage accumulation in the liver as well as increasing cytokine secretion and hepatic glucose production. Moreover, VDR deletion increases atherosclerosis by enabling lipid-laden M2 monocytes to adhere, migrate, and carry cholesterol into the atherosclerotic plaque and by increasing macrophage cholesterol uptake and esterification. Increased foam cell formation results from lack of VDR-SERCA2b interaction, causing SERCA dysfunction, activation of ER stress-CaMKII-JNKp-PPARγ signaling, and induction of the scavenger receptors CD36 and SR-A1. Bone marrow transplant of VDR-expressing cells into KODMAC mice improved insulin sensitivity, suppressed atherosclerosis, and decreased foam cell formation. The immunomodulatory effects of vitamin D in macrophages are thus critical in diet-induced insulin resistance and atherosclerosis in mice.

  9. Myelin-associated proteins labelled by slow axonal transport

    International Nuclear Information System (INIS)

    Giorgi, P.P.; DuBois, H.

    1981-01-01

    This paper deals with the problem of protein metabolism and provides evidence that the neuronal contribution to myelin metabolism may be restricted to lipids only. On the other hand this line of research led to the partial characterization of a group of neuronal proteins probably involved in axo-glial interactions subserving the onset of myelination and the structural maintenance of the mature myelin sheath. Intraocular injection of radioactive amino acids allows the study of the anterograde transport of labelled proteins along retinofugal fibres which are well myelinated. Myelin extracted from the optic nerve and tract under these conditions also contains labelled proteins. Three hypotheses are available to explain this phenomenon. To offer an explanation for this phenomenon the work was planned as follows. a) Characterization of the spatio-temporal pattern of labelling of myelin, in order to define the experimental conditions (survival time and region of the optic pathway to be studied) necessary to obtain maximal labelling. b) Characterization (by gel electrophoresis) of the myelin-associated proteins which become labelled by axonal transport, in order to work on a consistent pattern of labelling. c) Investigation of the possible mechanism responsible for the labelling of myelin-associated proteins. (Auth.)

  10. Convective-diffusive transport in protein crystal growth

    Science.gov (United States)

    Lin, H.; Rosenberger, F.; Alexander, J. I. D.; Nadarajah, A.

    1995-05-01

    Particular interest in the role of convection in protein crystallization has arisen since some protein single crystals of improved structural quality have been obtained under reduced gravity conditions. We have numerically modeled the time-dependent diffusive-convective transport in an isothermal protein crystal growth system at standard and zero gravity (1 g and 0 g). In the 2D model used, a rectangular crystal of fixed dimensions 400 μm × 600 μm is positioned at the bottom of a 1 mm high and 6 mm wide growth cell. The aqueous solution contains protein and precipitant. For the dependence of the crystal growth rate on interfacial supersaturation, experimental data for lysozyme are used. The repartitioning of water and precipitant at the growing interface is based on experimental segregation data for lysozyme: NaCl, and on complete rejection for a fictitious system in which lysozyme and precipitant have the same diffusivity. The results show that even in the small cell employed, protein concentration nonuniformities and gravity-driven solutal convection can be significant. The calculated convection velocities are of the same order of magnitude as those found in earlier experiments. As expected, convective transport enhances the growth rates. However, even when diffusion dominates mass transport, i.e. at 0 g, lysozyme crystal growth remains kinetically limited. Irrespective of the diffusivity of the precipitant, due to the low growth rates, the precipitant distribution in the solution remains rather uniform even at 0 g, unless strong coupling between precipitant and protein fluxes is assumed. The salt distribution in the crystal is predicted to be non-uniform at both 1 g and 0 g, as a consequence of protein depletion in the solution.

  11. HDL: The "Good" Cholesterol

    Science.gov (United States)

    ... There are two main types of cholesterol: HDL (good) cholesterol and LDL (bad) cholesterol: HDL stands for high-density lipoproteins. It is called the "good" cholesterol because it carries cholesterol from other parts ...

  12. What Is Cholesterol?

    Science.gov (United States)

    ... of Cholesterol There are two main types of cholesterol: LDL and HDL. The cholesterol blood test tells how much of each kind you have. Most cholesterol is LDL (low-density lipoprotein) cholesterol. This type is most ...

  13. Expression and putative role of mitochondrial transport proteins in cancer.

    Science.gov (United States)

    Lytovchenko, Oleksandr; Kunji, Edmund R S

    2017-08-01

    Cancer cells undergo major changes in energy and biosynthetic metabolism. One of them is the Warburg effect, in which pyruvate is used for fermentation rather for oxidative phosphorylation. Another major one is their increased reliance on glutamine, which helps to replenish the pool of Krebs cycle metabolites used for other purposes, such as amino acid or lipid biosynthesis. Mitochondria are central to these alterations, as the biochemical pathways linking these processes run through these organelles. Two membranes, an outer and inner membrane, surround mitochondria, the latter being impermeable to most organic compounds. Therefore, a large number of transport proteins are needed to link the biochemical pathways of the cytosol and mitochondrial matrix. Since the transport steps are relatively slow, it is expected that many of these transport steps are altered when cells become cancerous. In this review, changes in expression and regulation of these transport proteins are discussed as well as the role of the transported substrates. This article is part of a Special Issue entitled Mitochondria in Cancer, edited by Giuseppe Gasparre, Rodrigue Rossignol and Pierre Sonveaux. Copyright © 2017. Published by Elsevier B.V.

  14. The association of 83 plasma proteins with CHD mortality, BMI, HDL-, and total-cholesterol in men: applying multivariate statistics to identify proteins with prognostic value and biological relevance.

    Science.gov (United States)

    Heidema, A Geert; Thissen, Uwe; Boer, Jolanda M A; Bouwman, Freek G; Feskens, Edith J M; Mariman, Edwin C M

    2009-06-01

    In this study, we applied the multivariate statistical tool Partial Least Squares (PLS) to analyze the relative importance of 83 plasma proteins in relation to coronary heart disease (CHD) mortality and the intermediate end points body mass index, HDL-cholesterol and total cholesterol. From a Dutch monitoring project for cardiovascular disease risk factors, men who died of CHD between initial participation (1987-1991) and end of follow-up (January 1, 2000) (N = 44) and matched controls (N = 44) were selected. Baseline plasma concentrations of proteins were measured by a multiplex immunoassay. With the use of PLS, we identified 15 proteins with prognostic value for CHD mortality and sets of proteins associated with the intermediate end points. Subsequently, sets of proteins and intermediate end points were analyzed together by Principal Components Analysis, indicating that proteins involved in inflammation explained most of the variance, followed by proteins involved in metabolism and proteins associated with total-C. This study is one of the first in which the association of a large number of plasma proteins with CHD mortality and intermediate end points is investigated by applying multivariate statistics, providing insight in the relationships among proteins, intermediate end points and CHD mortality, and a set of proteins with prognostic value.

  15. Modelling Transcapillary Transport of Fluid and Proteins in Hemodialysis Patients.

    Directory of Open Access Journals (Sweden)

    Mauro Pietribiasi

    Full Text Available The kinetics of protein transport to and from the vascular compartment play a major role in the determination of fluid balance and plasma refilling during hemodialysis (HD sessions. In this study we propose a whole-body mathematical model describing water and protein shifts across the capillary membrane during HD and compare its output to clinical data while evaluating the impact of choosing specific values for selected parameters.The model follows a two-compartment structure (vascular and interstitial space and is based on balance equations of protein mass and water volume in each compartment. The capillary membrane was described according to the three-pore theory. Two transport parameters, the fractional contribution of large pores (αLP and the total hydraulic conductivity (LpS of the capillary membrane, were estimated from patient data. Changes in the intensity and direction of individual fluid and solute flows through each part of the transport system were analyzed in relation to the choice of different values of small pores radius and fractional conductivity, lymphatic sensitivity to hydraulic pressure, and steady-state interstitial-to-plasma protein concentration ratio.The estimated values of LpS and αLP were respectively 10.0 ± 8.4 mL/min/mmHg (mean ± standard deviation and 0.062 ± 0.041. The model was able to predict with good accuracy the profiles of plasma volume and serum total protein concentration in most of the patients (average root-mean-square deviation < 2% of the measured value.The applied model provides a mechanistic interpretation of fluid transport processes induced by ultrafiltration during HD, using a minimum of tuned parameters and assumptions. The simulated values of individual flows through each kind of pore and lymphatic absorption rate yielded by the model may suggest answers to unsolved questions on the relative impact of these not-measurable quantities on total vascular refilling and fluid balance.

  16. Exploitation of an iron transporter for bacterial protein antibiotic import.

    Science.gov (United States)

    White, Paul; Joshi, Amar; Rassam, Patrice; Housden, Nicholas G; Kaminska, Renata; Goult, Jonathan D; Redfield, Christina; McCaughey, Laura C; Walker, Daniel; Mohammed, Shabaz; Kleanthous, Colin

    2017-11-07

    Unlike their descendants, mitochondria and plastids, bacteria do not have dedicated protein import systems. However, paradoxically, import of protein bacteriocins, the mechanisms of which are poorly understood, underpins competition among pathogenic and commensal bacteria alike. Here, using X-ray crystallography, isothermal titration calorimetry, confocal fluorescence microscopy, and in vivo photoactivatable cross-linking of stalled translocation intermediates, we demonstrate how the iron transporter FpvAI in the opportunistic pathogen Pseudomonas aeruginosa is hijacked to translocate the bacteriocin pyocin S2 (pyoS2) across the outer membrane (OM). FpvAI is a TonB-dependent transporter (TBDT) that actively imports the small siderophore ferripyoverdine (Fe-Pvd) by coupling to the proton motive force (PMF) via the inner membrane (IM) protein TonB1. The crystal structure of the N-terminal domain of pyoS2 (pyoS2 NTD ) bound to FpvAI ( K d = 240 pM) reveals that the pyocin mimics Fe-Pvd, inducing the same conformational changes in the receptor. Mimicry leads to fluorescently labeled pyoS2 NTD being imported into FpvAI-expressing P. aeruginosa cells by a process analogous to that used by bona fide TBDT ligands. PyoS2 NTD induces unfolding by TonB1 of a force-labile portion of the plug domain that normally occludes the central channel of FpvAI. The pyocin is then dragged through this narrow channel following delivery of its own TonB1-binding epitope to the periplasm. Hence, energized nutrient transporters in bacteria also serve as rudimentary protein import systems, which, in the case of FpvAI, results in a protein antibiotic 60-fold bigger than the transporter's natural substrate being translocated across the OM. Copyright © 2017 the Author(s). Published by PNAS.

  17. The effect of simvastatin treatment on the amyloid precursor protein and brain cholesterol metabolism in patients with Alzheimer's disease

    DEFF Research Database (Denmark)

    Hoglund, K; Thelen, K M; Syversen, S

    2005-01-01

    as on cognitive decline in patients with AD. Despite biochemical data suggesting that treatment with 20 mg/day of simvastatin for 12 months does affect the brain cholesterol metabolism, we did not find any change in CSF or plasma levels of beta-amyloid (Abeta)(1-42). However, by analysis of APP isoforms, we found...... with AD have been treated with simvastatin (20 mg/day) for 12 months. The aim was to further investigate the effect of simvastatin treatment on cerebrospinal fluid (CSF) biomarkers of APP processing, AD biomarkers as total tau and tau phosphorylated at threonine 181, brain cholesterol metabolism as well...

  18. STARD4 knockdown in HepG2 cells disrupts cholesterol trafficking associated with the plasma membrane, ER, and ERC.

    Science.gov (United States)

    Garbarino, Jeanne; Pan, Meihui; Chin, Harvey F; Lund, Frederik W; Maxfield, Frederick R; Breslow, Jan L

    2012-12-01

    STARD4, a member of the evolutionarily conserved START gene family, has been implicated in the nonvesicular intracellular transport of cholesterol. However, the direction of transport and the membranes with which this protein interacts are not clear. We present studies of STARD4 function using small hairpin RNA knockdown technology to reduce STARD4 expression in HepG2 cells. In a cholesterol-poor environment, we found that a reduction in STARD4 expression leads to retention of cholesterol at the plasma membrane, reduction of endoplasmic reticulum-associated cholesterol, and decreased ACAT synthesized cholesteryl esters. Furthermore, D4 KD cells exhibited a reduced rate of sterol transport to the endocytic recycling compartment after cholesterol repletion. Although these cells displayed normal endocytic trafficking in cholesterol-poor and replete conditions, cell surface low density lipoprotein receptor (LDLR) levels were increased and decreased, respectively. We also observed a decrease in NPC1 protein expression, suggesting the induction of compensatory pathways to maintain cholesterol balance. These data indicate a role for STARD4 in nonvesicular transport of cholesterol from the plasma membrane and the endocytic recycling compartment to the endoplasmic reticulum and perhaps other intracellular compartments as well.

  19. Intestinal cholesterol secretion: future clinical implications

    NARCIS (Netherlands)

    Jakulj, L.; Besseling, J.; Stroes, E. S. G.; Groen, A. K.

    2013-01-01

    Together with the liver, the intestine serves as a homeostatic organ in cholesterol metabolism. Recent evidence has substantiated the pivotal role of the intestine in reverse cholesterol transport (RCT). RCT is a fundamental antiatherogenic pathway, mediating the removal of cholesterol from tissues

  20. Intestinal cholesterol secretion : future clinical implications

    NARCIS (Netherlands)

    Jakulj, L.; Besseling, J.; Stroes, E. S. G.; Groen, A. K.

    2013-01-01

    Together with the liver, the intestine serves as a homeostatic organ in cholesterol metabolism. Recent evidence has substantiated the pivotal role of the intestine in reverse cholesterol transport (RCT). RCT is a fundamental antiatherogenic pathway, mediating the removal of cholesterol from tissues

  1. Sensitivity to lysosome-dependent cell death is directly regulated by lysosomal cholesterol content.

    Directory of Open Access Journals (Sweden)

    Hanna Appelqvist

    Full Text Available Alterations in lipid homeostasis are implicated in several neurodegenerative diseases, although the mechanisms responsible are poorly understood. We evaluated the impact of cholesterol accumulation, induced by U18666A, quinacrine or mutations in the cholesterol transporting Niemann-Pick disease type C1 (NPC1 protein, on lysosomal stability and sensitivity to lysosome-mediated cell death. We found that neurons with lysosomal cholesterol accumulation were protected from oxidative stress-induced apoptosis. In addition, human fibroblasts with cholesterol-loaded lysosomes showed higher lysosomal membrane stability than controls. Previous studies have shown that cholesterol accumulation is accompanied by the storage of lipids such as sphingomyelin, glycosphingolipids and sphingosine and an up regulation of lysosomal associated membrane protein-2 (LAMP-2, which may also influence lysosomal stability. However, in this study the use of myriocin and LAMP deficient fibroblasts excluded these factors as responsible for the rescuing effect and instead suggested that primarily lysosomal cholesterol content determineD the cellular sensitivity to toxic insults. Further strengthening this concept, depletion of cholesterol using methyl-β-cyclodextrin or 25-hydroxycholesterol decreased the stability of lysosomes and cells became more prone to undergo apoptosis. In conclusion, cholesterol content regulated lysosomal membrane permeabilization and thereby influenced cell death sensitivity. Our data suggests that lysosomal cholesterol modulation might be used as a therapeutic strategy for conditions associated with accelerated or repressed apoptosis.

  2. Nanoscale Electron Transport Measurements of Immobilized Cytochrome P450 Proteins

    Science.gov (United States)

    Bostick, Christopher D.; Flora, Darcy R.; Gannett, Peter M.; Tracy, Timothy S.; Lederman, David

    2015-01-01

    Gold nanopillars, functionalized with an organic self-assembled monolayer, can be used to measure the electrical conductance properties of immobilized proteins without aggregation. Measurements of the conductance of nanopillars with cytochrome P450 2C9 (CYP2C9) proteins using conducting probe atomic force microscopy demonstrate that a correlation exists between the energy barrier height between hopping sites and CYP2C9 metabolic activity. Measurements performed as a function of tip force indicate that, when subjected to a large force, the protein is more stable in the presence of a substrate. This agrees with the hypothesis that substrate entry into the active site helps to stabilize the enzyme. The relative distance between hopping sites also increases with increasing force, possibly because protein functional groups responsible for electron transport depend on the structure of the protein. The inhibitor sulfaphenazole, in addition to the previously studied aniline, increased the barrier height for electron transfer and thereby makes CYP2C9 reduction more difficult and inhibits metabolism. This suggests that P450 Type II binders may decrease the ease of electron transport processes in the enzyme, in addition to occupying the active site. PMID:25804257

  3. Transport of Proteins Dissolved in Organic Solvents Across Biomimetic Membranes

    Science.gov (United States)

    Bromberg, Lev E.; Klibanov, Alexander M.

    1995-02-01

    Using lipid-impregnated porous cellulose membranes as biomimetic barriers, we tested the hypothesis that to afford effective transmembrane transfer of proteins and nucleic acids, the vehicle solvent should be able to dissolve both the biopolymers and the lipids. While the majority of solvents dissolve one or the other, ethanol and methanol were found to dissolve both, especially if the protein had been lyophilized from an aqueous solution of a pH remote from the protein's isoelectric point. A number of proteins, as well as RNA and DNA, dissolved in these alcohols readily crossed the lipidized membranes, whereas the same biopolymers placed in nondissolving solvents (e.g., hexane and ethyl acetate) or in those unable to dissolve lipids (e.g., water and dimethyl sulfoxide) exhibited little transmembrane transport. The solubility of biopolymers in ethanol and methanol was further enhanced by complexation with detergents and poly(ethylene glycol); significant protein and nucleic acid transport through the lipidized membranes was observed from these solvents but not from water.

  4. Amphipathic alpha-helices and putative cholesterol binding domains of the influenza virus matrix M1 protein are crucial for virion structure organisation.

    Science.gov (United States)

    Tsfasman, Tatyana; Kost, Vladimir; Markushin, Stanislav; Lotte, Vera; Koptiaeva, Irina; Bogacheva, Elena; Baratova, Ludmila; Radyukhin, Victor

    2015-12-02

    The influenza virus matrix M1 protein is an amphitropic membrane-associated protein, forming the matrix layer immediately beneath the virus raft membrane, thereby ensuring the proper structure of the influenza virion. The objective of this study was to elucidate M1 fine structural characteristics, which determine amphitropic properties and raft membrane activities of the protein, via 3D in silico modelling with subsequent mutational analysis. Computer simulations suggest the amphipathic nature of the M1 α-helices and the existence of putative cholesterol binding (CRAC) motifs on six amphipathic α-helices. Our finding explains for the first time many features of this protein, particularly the amphitropic properties and raft/cholesterol binding potential. To verify these results, we generated mutants of the A/WSN/33 strain via reverse genetics. The M1 mutations included F32Y in the CRAC of α-helix 2, W45Y and W45F in the CRAC of α-helix 3, Y100S in the CRAC of α-helix 6, M128A and M128S in the CRAC of α-helix 8 and a double L103I/L130I mutation in both a putative cholesterol consensus motif and the nuclear localisation signal. All mutations resulted in viruses with unusual filamentous morphology. Previous experimental data regarding the morphology of M1-gene mutant influenza viruses can now be explained in structural terms and are consistent with the pivotal role of the CRAC-domains and amphipathic α-helices in M1-lipid interactions. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Cholesterol depletion of hepatoma cells impairs hepatitis B virus envelopment by altering the topology of the large envelope protein.

    NARCIS (Netherlands)

    Dorobantu, C.M.; Macovei, A.; Lazar, C.; Dwek, R.A.; Zitzmann, N.; Branza-Nichita, N.

    2011-01-01

    Previous reports have shown that cholesterol depletion of the membrane envelope of the hepatitis B virus (HBV) impairs viral infection of target cells. A potential function of this lipid in later steps of the viral life cycle remained controversial, with secretion of virions and subviral particles

  6. Analysis of Cholesterol Trafficking with Fluorescent Probes

    DEFF Research Database (Denmark)

    Maxfield, Frederick R.; Wustner, Daniel

    2012-01-01

    Cholesterol plays an important role in determining the biophysical properties of biological membranes, and its concentration is tightly controlled by homeostatic processes. The intracellular transport of cholesterol among organelles is a key part of the homeostatic mechanism, but sterol transport...... that can bind to cholesterol to reveal its distribution in cells. We also discuss the use of intrinsically fluorescent sterols that closely mimic cholesterol, as well as some minimally modified fluorophore-labeled sterols. Methods for imaging these sterols by conventional fluorescence microscopy...... and by multiphoton microscopy are described. Some label-free methods for imaging cholesterol itself are also discussed briefly....

  7. The PIN-FORMED (PIN) protein family of auxin transporters

    Czech Academy of Sciences Publication Activity Database

    Křeček, Pavel; Skůpa, Petr; Libus, Jiří; Naramoto, S.; Tejos, R.; Friml, J.; Zažímalová, Eva

    2009-01-01

    Roč. 10, č. 12 (2009), s. 249.1-249.11 ISSN 1474-760X R&D Projects: GA MŠk(CZ) LC06034; GA AV ČR KJB600380904; GA AV ČR(CZ) IAA601630703 Institutional research plan: CEZ:AV0Z50380511 Keywords : PIN protein family * auxin efflux carriers * auxin transport Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.626, year: 2009

  8. Loss of Subcellular Lipid Transport Due to ARV1 Deficiency Disrupts Organelle Homeostasis and Activates the Unfolded Protein Response*

    Science.gov (United States)

    Shechtman, Caryn F.; Henneberry, Annette L.; Seimon, Tracie A.; Tinkelenberg, Arthur H.; Wilcox, Lisa J.; Lee, Eunjee; Fazlollahi, Mina; Munkacsi, Andrew B.; Bussemaker, Harmen J.; Tabas, Ira; Sturley, Stephen L.

    2011-01-01

    The ARV1-encoded protein mediates sterol transport from the endoplasmic reticulum (ER) to the plasma membrane. Yeast ARV1 mutants accumulate multiple lipids in the ER and are sensitive to pharmacological modulators of both sterol and sphingolipid metabolism. Using fluorescent and electron microscopy, we demonstrate sterol accumulation, subcellular membrane expansion, elevated lipid droplet formation, and vacuolar fragmentation in ARV1 mutants. Motif-based regression analysis of ARV1 deletion transcription profiles indicates activation of Hac1p, an integral component of the unfolded protein response (UPR). Accordingly, we show constitutive splicing of HAC1 transcripts, induction of a UPR reporter, and elevated expression of UPR targets in ARV1 mutants. IRE1, encoding the unfolded protein sensor in the ER lumen, exhibits a lethal genetic interaction with ARV1, indicating a viability requirement for the UPR in cells lacking ARV1. Surprisingly, ARV1 mutants expressing a variant of Ire1p defective in sensing unfolded proteins are viable. Moreover, these strains also exhibit constitutive HAC1 splicing that interacts with DTT-mediated perturbation of protein folding. These data suggest that a component of UPR induction in arv1Δ strains is distinct from protein misfolding. Decreased ARV1 expression in murine macrophages also results in UPR induction, particularly up-regulation of activating transcription factor-4, CHOP (C/EBP homologous protein), and apoptosis. Cholesterol loading or inhibition of cholesterol esterification further elevated CHOP expression in ARV1 knockdown cells. Thus, loss or down-regulation of ARV1 disturbs membrane and lipid homeostasis, resulting in a disruption of ER integrity, one consequence of which is induction of the UPR. PMID:21266578

  9. Diets containing soy or rice protein isolate increase insulin sensitivity and improve lipid homeostasis in weanling rats fed high fat, high cholesterol Western diets as a result of activation of PPAR and LXR-mediated pathways

    Science.gov (United States)

    The current study examined the effects of feeding soy protein isolate (SPI) and rice protein isolate (RPI) on insulin sensitivity and fat breakdown in weanling rats consuming high fat/high cholesterol diets. Male Sprague-Dawley rats were placed on semi-purified diets containing the milk protein case...

  10. Hydrophobic interaction chromatography of proteins. IV. Protein adsorption capacity and transport in preparative mode.

    Science.gov (United States)

    To, Brian C S; Lenhoff, Abraham M

    2011-01-21

    The adsorption isotherms of four model proteins (lysozyme, α-lactalbumin, ovalbumin, and BSA) on eight commercial phenyl hydrophobic interaction chromatography media were measured. The isotherms were softer than those usually seen in ion-exchange chromatography of proteins, and the static capacities of the media were lower, ranging from 30 to 110 mg/mL, depending on the ammonium sulfate concentration and the protein and adsorbent types. The protein-accessible surface area appears to be the main factor determining the binding capacity, and little correlation was seen with the protein affinities of the adsorbents. Breakthrough experiments showed that the dynamic capacities of the adsorbents at 10% breakthrough were 20-80% of the static capacities, depending on adsorbent type. Protein diffusivities in the adsorbents were estimated from batch uptake experiments using the pore diffusion and homogeneous diffusion models. Protein transport was affected by the adsorbent pore structures. Apparent diffusivities were higher at lower salt concentrations and column loadings, suggesting that adsorbed proteins may retard intraparticle protein transport. The diffusivities estimated from the batch uptake experiments were used to predict column breakthrough behavior. Analytical solutions developed for ion-exchange systems were able to provide accurate predictions for lysozyme breakthrough but not for ovalbumin. Impurities in the ovalbumin solutions used for the breakthrough experiments may have affected the ovalbumin uptake and led to the discrepancies between the predictions and the experimental results. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. JTT-130, a microsomal triglyceride transfer protein (MTP inhibitor lowers plasma triglycerides and LDL cholesterol concentrations without increasing hepatic triglycerides in guinea pigs

    Directory of Open Access Journals (Sweden)

    Shrestha Sudeep

    2005-09-01

    Full Text Available Abstract Background Microsomal transfer protein inhibitors (MTPi have the potential to be used as a drug to lower plasma lipids, mainly plasma triglycerides (TG. However, studies with animal models have indicated that MTPi treatment results in the accumulation of hepatic TG. The purpose of this study was to evaluate whether JTT-130, a unique MTPi, targeted to the intestine, would effectively reduce plasma lipids without inducing a fatty liver. Methods Male guinea pigs (n = 10 per group were used for this experiment. Initially all guinea pigs were fed a hypercholesterolemic diet containing 0.08 g/100 g dietary cholesterol for 3 wk. After this period, animals were randomly assigned to diets containing 0 (control, 0.0005 or 0.0015 g/100 g of MTPi for 4 wk. A diet containing 0.05 g/100 g of atorvastatin, an HMG-CoA reductase inhibitor was used as the positive control. At the end of the 7th week, guinea pigs were sacrificed to assess drug effects on plasma and hepatic lipids, composition of LDL and VLDL, hepatic cholesterol and lipoprotein metabolism. Results Plasma LDL cholesterol and TG were 25 and 30% lower in guinea pigs treated with MTPi compared to controls (P Conclusion These results suggest that JTT-130 could have potential clinical applications due to its plasma lipid lowering effects with no alterations in hepatic lipid concentrations.

  12. Circulating levels of linoleic acid and HDL-cholesterol are major determinants of 4-hydroxynonenal protein adducts in patients with heart failure☆

    Science.gov (United States)

    Asselin, Caroline; Ducharme, Anique; Ntimbane, Thierry; Ruiz, Matthieu; Fortier, Annik; Guertin, Marie-Claude; Lavoie, Joël; Diaz, Ariel; Levy, Émile; Tardif, Jean-Claude; Des Rosiers, Christine

    2013-01-01

    Objective Measurements of oxidative stress biomarkers in patients with heart failure (HF) have yielded controversial results. This study aimed at testing the hypothesis that circulating levels of the lipid peroxidation product 4-hydroxynonenal bound to thiol proteins (4HNE-P) are strongly associated with those of its potential precursors, namely n-6 polyunsaturated fatty acids (PUFA). Methods and results Circulating levels of 4HNE-P were evaluated by gas chromatography-mass spectrometry in 71 control subjects and 61 ambulatory symptomatic HF patients along with various other clinically- and biochemically-relevant parameters, including other oxidative stress markers, and total levels of fatty acids from all classes, which reflect both free and bound to cholesterol, phospholipids and triglycerides. All HF patients had severe systolic functional impairment despite receiving optimal evidence-based therapies. Compared to controls, HF patients displayed markedly lower circulating levels of HDL- and LDL-cholesterol, which are major PUFA carriers, as well as of PUFA of the n-6 series, specifically linoleic acid (LA; P=0.001). Circulating 4HNE-P in HF patients was similar to controls, albeit multiple regression analysis revealed that LA was the only factor that was significantly associated with circulating 4HNE-P in the entire population (R2=0.086; P=0.02). In HF patients only, 4HNE-P was even more strongly associated with LA (P=0.003) and HDL-cholesterol (pHF group only. Conclusion Results from this study emphasize the importance of considering changes in lipids and lipoproteins in the interpretation of measurements of lipid peroxidation products. Further studies appear warranted to explore the possibility that HDL-cholesterol particles may be a carrier of 4HNE adducts. PMID:24494189

  13. Circulating levels of linoleic acid and HDL-cholesterol are major determinants of 4-hydroxynonenal protein adducts in patients with heart failure.

    Science.gov (United States)

    Asselin, Caroline; Ducharme, Anique; Ntimbane, Thierry; Ruiz, Matthieu; Fortier, Annik; Guertin, Marie-Claude; Lavoie, Joël; Diaz, Ariel; Levy, Emile; Tardif, Jean-Claude; Des Rosiers, Christine

    2014-01-01

    Measurements of oxidative stress biomarkers in patients with heart failure (HF) have yielded controversial results. This study aimed at testing the hypothesis that circulating levels of the lipid peroxidation product 4-hydroxynonenal bound to thiol proteins (4HNE-P) are strongly associated with those of its potential precursors, namely n-6 polyunsaturated fatty acids (PUFA). Circulating levels of 4HNE-P were evaluated by gas chromatography-mass spectrometry in 71 control subjects and 61 ambulatory symptomatic HF patients along with various other clinically- and biochemically-relevant parameters, including other oxidative stress markers, and total levels of fatty acids from all classes, which reflect both free and bound to cholesterol, phospholipids and triglycerides. All HF patients had severe systolic functional impairment despite receiving optimal evidence-based therapies. Compared to controls, HF patients displayed markedly lower circulating levels of HDL- and LDL-cholesterol, which are major PUFA carriers, as well as of PUFA of the n-6 series, specifically linoleic acid (LA; P=0.001). Circulating 4HNE-P in HF patients was similar to controls, albeit multiple regression analysis revealed that LA was the only factor that was significantly associated with circulating 4HNE-P in the entire population (R (2)=0.086; P=0.02). In HF patients only, 4HNE-P was even more strongly associated with LA (P=0.003) and HDL-cholesterol (pHF group only. Results from this study emphasize the importance of considering changes in lipids and lipoproteins in the interpretation of measurements of lipid peroxidation products. Further studies appear warranted to explore the possibility that HDL-cholesterol particles may be a carrier of 4HNE adducts.

  14. Effect of Atorvastatin on Serum Levels of Total Cholesterol and High-Sensitivity C-reactive Protein in High-Risk Patients with Atrial Fibrillation in Asia.

    Science.gov (United States)

    Shi, Ming Yu; Xue, Feng Hua; Teng, Shi Chao; Jiang, Li; Zhu, Jing; Yin, Feng; Gu, Hong Yue

    2015-08-01

    The aim of this meta-analysis was to investigate the effects of atorvastatin on serum levels of high-sensitivity C-reactive protein (hs-CRP) and total cholesterol in atrial fibrillation (AF) patients in Asia. By searching English and Chinese language-based electronic databases (ie, PubMed, EBSCO, Ovid, SpringerLink, Wiley, Web of Science, Wanfang database, China National Knowledge Infrastructure, and VIP database), we identified 13 studies relevant to our topic of interest. Data were collected from the 13 studies and analyzed with Comprehensive Meta-Analysis software (version 2.0, Biostat Inc., Englewood, New Jersey). Initially, our database searches retrieved 356 studies (45 in English, 311 in Chinese). Thirteen studies were selected for the meta-analysis following stringent criteria. The data included 1239 patients with AF, of whom 634 were treated with atorvastatin and included in the treatment group, and 605 patients were treated with conventional treatment and included in the control group. The results of our meta-analysis suggested that the serum levels of hs-CRP (mg/L) and total cholesterol (mmol/L) in the treatment group were significantly lower than those of the control group (hs-CRP: standardized mean difference = 0.962; 95% CI, 0.629-1.295, P < 0.001; total cholesterol: standardized mean difference = 1.400; 95% CI, 0.653-2.146, P < 0.001). The findings of this study suggest that atorvastatin may be very effective in decreasing serum levels of hs-CRP and total cholesterol to prevent cardiovascular events. Copyright © 2015 Elsevier HS Journals, Inc. All rights reserved.

  15. Beyond the Cholesterol-Lowering Effect of Soy Protein: A Review of the Effects of Dietary Soy and Its Constituents on Risk Factors for Cardiovascular Disease

    Directory of Open Access Journals (Sweden)

    D. Dan Ramdath

    2017-03-01

    Full Text Available The hypocholesterolemic effect of soy is well-documented and this has led to the regulatory approval of a health claim relating soy protein to a reduced risk of cardiovascular disease (CVD. However, soybeans contain additional components, such as isoflavones, lecithins, saponins and fiber that may improve cardiovascular health through independent mechanisms. This review summarizes the evidence on the cardiovascular benefits of non-protein soy components in relation to known CVD risk factors such as hypertension, hyperglycemia, inflammation, and obesity beyond cholesterol lowering. Overall, the available evidence suggests non-protein soy constituents improve markers of cardiovascular health; however, additional carefully designed studies are required to independently elucidate these effects. Further, work is also needed to clarify the role of isoflavone-metabolizing phenotype and gut microbiota composition on biological effect.

  16. Beyond the Cholesterol-Lowering Effect of Soy Protein: A Review of the Effects of Dietary Soy and Its Constituents on Risk Factors for Cardiovascular Disease.

    Science.gov (United States)

    Ramdath, D Dan; Padhi, Emily M T; Sarfaraz, Sidra; Renwick, Simone; Duncan, Alison M

    2017-03-24

    The hypocholesterolemic effect of soy is well-documented and this has led to the regulatory approval of a health claim relating soy protein to a reduced risk of cardiovascular disease (CVD). However, soybeans contain additional components, such as isoflavones, lecithins, saponins and fiber that may improve cardiovascular health through independent mechanisms. This review summarizes the evidence on the cardiovascular benefits of non-protein soy components in relation to known CVD risk factors such as hypertension, hyperglycemia, inflammation, and obesity beyond cholesterol lowering. Overall, the available evidence suggests non-protein soy constituents improve markers of cardiovascular health; however, additional carefully designed studies are required to independently elucidate these effects. Further, work is also needed to clarify the role of isoflavone-metabolizing phenotype and gut microbiota composition on biological effect.

  17. LRP1 controls cPLA2 phosphorylation, ABCA1 expression and cellular cholesterol export.

    Directory of Open Access Journals (Sweden)

    Li Zhou

    2009-08-01

    Full Text Available ATP-binding cassette transporter A1 mediates apolipoprotein AI-dependent efflux of cholesterol and thereby removes cholesterol from peripheral tissues. ABCA1 expression is tightly regulated and deficiency of this cholesterol transporter results in cholesterol accumulation within cells. Low-density lipoprotein receptor-related protein 1 (LRP1 participates in lipid metabolism and energy homeostasis by endocytosis of apolipoprotein E-containing lipoproteins and modulation of cellular proliferation signals.In the present study, we demonstrate a new role for LRP1 in reverse cholesterol transport. Absence of LRP1 expression results in increased PDGFRbeta signaling and sequential activation of the mitogen-activated protein kinase signaling pathway, which increases phosphorylation of cytosolic phospholipase A(2 (cPLA(2. Phosphorylated and activated cPLA(2 releases arachidonic acid from the phospholipid pool. Overproduction of arachidonic acid suppresses the activation of LXR/RXR heterodimers bound to the promoter of LXR regulated genes such as ABCA1, resulting in greatly reduced ABCA1 expression.LRP1 regulates LXR-mediated gene transcription and participates in reverse cholesterol transport by controlling cPLA(2 activation and ABCA1 expression. LRP1 thus functions as a physiological integrator of cellular lipid homeostasis with signals that regulate cellular proliferation and vascular wall integrity.

  18. Chloroplast Iron Transport Proteins - Function and Impact on Plant Physiology.

    Science.gov (United States)

    López-Millán, Ana F; Duy, Daniela; Philippar, Katrin

    2016-01-01

    Chloroplasts originated about three billion years ago by endosymbiosis of an ancestor of today's cyanobacteria with a mitochondria-containing host cell. During evolution chloroplasts of higher plants established as the site for photosynthesis and thus became the basis for all life dependent on oxygen and carbohydrate supply. To fulfill this task, plastid organelles are loaded with the transition metals iron, copper, and manganese, which due to their redox properties are essential for photosynthetic electron transport. In consequence, chloroplasts for example represent the iron-richest system in plant cells. However, improvement of oxygenic photosynthesis in turn required adaptation of metal transport and homeostasis since metal-catalyzed generation of reactive oxygen species (ROS) causes oxidative damage. This is most acute in chloroplasts, where radicals and transition metals are side by side and ROS-production is a usual feature of photosynthetic electron transport. Thus, on the one hand when bound by proteins, chloroplast-intrinsic metals are a prerequisite for photoautotrophic life, but on the other hand become toxic when present in their highly reactive, radical generating, free ionic forms. In consequence, transport, storage and cofactor-assembly of metal ions in plastids have to be tightly controlled and are crucial throughout plant growth and development. In the recent years, proteins for iron transport have been isolated from chloroplast envelope membranes. Here, we discuss their putative functions and impact on cellular metal homeostasis as well as photosynthetic performance and plant metabolism. We further consider the potential of proteomic analyses to identify new players in the field.

  19. Lipid-protein interactions. The leucine transport system of Lactococcus lactis.

    NARCIS (Netherlands)

    Veld, Geertruida Elisabeth in 't

    1992-01-01

    In summary, it is concluded, that a functionally reconstituted leucine transport system of L. lactis is affected by bilayer features in the following order of importance: lipid headgroup (H+-bonding) › acyl chain carbon number (thickness) › cholesterol (fluidity) › acyl chain unsaturation (indirect

  20. DISP3, a sterol-sensing domain-containing protein that links thyroid hormone action and cholesterol metabolism

    Czech Academy of Sciences Publication Activity Database

    Zíková, Martina; Corlett, Alicia; Bendová, Zdeňka; Pajer, Petr; Bartůněk, Petr

    2009-01-01

    Roč. 23, č. 4 (2009), s. 520-528 ISSN 0888-8809 R&D Projects: GA AV ČR IAA500520705 Grant - others:EC(XE) LSHM-CT-2005-018652 Institutional research plan: CEZ:AV0Z50520514; CEZ:AV0Z50110509 Keywords : thyroid hormone receptor * cholesterol metabolism * sterol-sensing domain Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.257, year: 2009

  1. Soy-dairy protein blend and whey protein ingestion after resistance exercise increases amino acid transport and transporter expression in human skeletal muscle

    Science.gov (United States)

    Reidy, P. T.; Walker, D. K.; Dickinson, J. M.; Gundermann, D. M.; Drummond, M. J.; Timmerman, K. L.; Cope, M. B.; Mukherjea, R.; Jennings, K.; Volpi, E.

    2014-01-01

    Increasing amino acid availability (via infusion or ingestion) at rest or postexercise enhances amino acid transport into human skeletal muscle. It is unknown whether alterations in amino acid availability, from ingesting different dietary proteins, can enhance amino acid transport rates and amino acid transporter (AAT) mRNA expression. We hypothesized that the prolonged hyperaminoacidemia from ingesting a blend of proteins with different digestion rates postexercise would enhance amino acid transport into muscle and AAT expression compared with the ingestion of a rapidly digested protein. In a double-blind, randomized clinical trial, we studied 16 young adults at rest and after acute resistance exercise coupled with postexercise (1 h) ingestion of either a (soy-dairy) protein blend or whey protein. Phenylalanine net balance and transport rate into skeletal muscle were measured using stable isotopic methods in combination with femoral arteriovenous blood sampling and muscle biopsies obtained at rest and 3 and 5 h postexercise. Phenylalanine transport into muscle and mRNA expression of select AATs [system L amino acid transporter 1/solute-linked carrier (SLC) 7A5, CD98/SLC3A2, system A amino acid transporter 2/SLC38A2, proton-assisted amino acid transporter 1/SLC36A1, cationic amino acid transporter 1/SLC7A1] increased to a similar extent in both groups (P protein blend resulted in a prolonged and positive net phenylalanine balance during postexercise recovery compared with whey protein (P protein synthesis increased similarly between groups. We conclude that, while both protein sources enhanced postexercise AAT expression, transport into muscle, and myofibrillar protein synthesis, postexercise ingestion of a protein blend results in a slightly prolonged net amino acid balance across the leg compared with whey protein. PMID:24699854

  2. Photoaffinity labeling with cholesterol analogues precisely maps a cholesterol-binding site in voltage-dependent anion channel-1.

    Science.gov (United States)

    Budelier, Melissa M; Cheng, Wayland W L; Bergdoll, Lucie; Chen, Zi-Wei; Janetka, James W; Abramson, Jeff; Krishnan, Kathiresan; Mydock-McGrane, Laurel; Covey, Douglas F; Whitelegge, Julian P; Evers, Alex S

    2017-06-02

    Voltage-dependent anion channel-1 (VDAC1) is a highly regulated β-barrel membrane protein that mediates transport of ions and metabolites between the mitochondria and cytosol of the cell. VDAC1 co-purifies with cholesterol and is functionally regulated by cholesterol, among other endogenous lipids. Molecular modeling studies based on NMR observations have suggested five cholesterol-binding sites in VDAC1, but direct experimental evidence for these sites is lacking. Here, to determine the sites of cholesterol binding, we photolabeled purified mouse VDAC1 (mVDAC1) with photoactivatable cholesterol analogues and analyzed the photolabeled sites with both top-down mass spectrometry (MS), and bottom-up MS paired with a clickable, stable isotope-labeled tag, FLI -tag. Using cholesterol analogues with a diazirine in either the 7 position of the steroid ring (LKM38) or the aliphatic tail (KK174), we mapped a binding pocket in mVDAC1 localized to Thr 83 and Glu 73 , respectively. When Glu 73 was mutated to a glutamine, KK174 no longer photolabeled this residue, but instead labeled the nearby Tyr 62 within this same binding pocket. The combination of analytical strategies employed in this work permits detailed molecular mapping of a cholesterol-binding site in a protein, including an orientation of the sterol within the site. Our work raises the interesting possibility that cholesterol-mediated regulation of VDAC1 may be facilitated through a specific binding site at the functionally important Glu 73 residue. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Membrane ion transport in erythrocytes of salt hypertensive Dahl rats and their F2 hybrids: the importance of cholesterol

    Czech Academy of Sciences Publication Activity Database

    Vokurková, Martina; Dobešová, Zdenka; Kuneš, Jaroslav; Zicha, Josef

    2003-01-01

    Roč. 26, č. 5 (2003), s. 397-404 ISSN 0916-9636 R&D Projects: GA ČR GA305/00/1638; GA MŠk LN00A069 Institutional research plan: CEZ:AV0Z5011922 Keywords : ion transport * salt hypertension * plasma lipids Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 2.013, year: 2003

  4. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) alters the mRNA expression of critical genes associated with cholesterol metabolism, bile acid biosynthesis, and bile transport in rat liver: A microarray study

    International Nuclear Information System (INIS)

    Fletcher, Nick; Wahlstroem, David; Lundberg, Rebecca; Nilsson, Charlotte B.; Nilsson, Kerstin C.; Stockling, Kenneth; Hellmold, Heike; Hakansson, Helen

    2005-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent hepatotoxin that exerts its toxicity through binding to the aryl hydrocarbon receptor (AhR) and the subsequent induction or repression of gene transcription. In order to further identify novel genes and pathways that may be associated with TCDD-induced hepatotoxicity, we investigated gene changes in rat liver following exposure to single oral doses of TCDD. Male Sprague-Dawley rats were administered single doses of 0.4 μg/kg bw or 40 μg/kg bw TCDD and killed at 6 h, 24 h, or 7 days, for global analyses of gene expression. In general, low-dose TCDD exposure resulted in greater than 2-fold induction of genes coding for a battery of phase I and phase II metabolizing enzymes including CYP1A1, CYP1A2, NADPH quinone oxidoreductase, UGT1A6/7, and metallothionein 1. However, 0.4 μg/kg bw TCDD also altered the expression of Gadd45a and Cyclin D1, suggesting that even low-dose TCDD exposure can alter the expression of genes indicative of cellular stress or DNA damage and associated with cell cycle control. At the high-dose, widespread changes were observed for genes encoding cellular signaling proteins, cellular adhesion, cytoskeletal and membrane transport proteins as well as transcripts coding for lipid, carbohydrate and nitrogen metabolism. In addition, decreased expression of cytochrome P450 7A1, short heterodimer partner (SHP; gene designation nr0b2), farnesyl X receptor (FXR), Ntcp, and Slc21a5 (oatp2) were observed and confirmed by RT-PCR analyses in independent rat liver samples. Altered expression of these genes implies major deregulation of cholesterol metabolism and bile acid synthesis and transport. We suggest that these early and novel changes have the potential to contribute significantly to TCDD induced hepatotoxicity and hypercholesterolemia

  5. Cholesterol IQ Quiz

    Science.gov (United States)

    ... Peripheral Artery Disease Venous Thromboembolism Aortic Aneurysm More Cholesterol IQ Quiz Updated:Jul 5,2017 Begin the quiz Cholesterol • Home • About Cholesterol Introduction Atherosclerosis What Your Cholesterol ...

  6. High Blood Cholesterol

    Science.gov (United States)

    ... To Health Topics / High Blood Cholesterol High Blood Cholesterol Also known as Hypercholesterolemia High blood cholesterol is ... Lipid panel tests to check for healthy blood cholesterol levels Doctors use lipid panels to check whether ...

  7. Water-mediated energy transport and structure across a protein-protein interface

    Science.gov (United States)

    Leitner, David

    2010-03-01

    Water molecules embedded within proteins or at the interface between globules play a central role in folding and function. We discuss the influence of interfacial water molecules on energy transport and structure, specifically the role of water at the interface between the two globules of the homodimeric hemoglobin from Scapharca inaequivalvis, which binds oxygen cooperatively. We have studied the water-mediated energy transport in this protein with communication maps and nonequilibrium molecular simulations of energy flow, which reveal the disproportionate amount of energy carried by the water molecules, particularly across the interface, i.e., a larger thermal conductivity of the interfacial waters compared with other parts of the protein, promoting hydrogen bond rearrangements at the interface.

  8. The Effects of Intrauterine Malnutrition on Maternal-Fetal Cholesterol Transport and Fetal Lipid Synthesis in Mice

    NARCIS (Netherlands)

    van Meer, Hester; van Straten, Esther M. E.; Baller, Julius F. W.; van Dijk, Theo H.; Kuipers, Folkert; Verkade, Henkjan J.; Plosch, Torsten

    Intrauterine malnutrition is associated with increased susceptibility to chronic diseases in adulthood. Growth-restricted infants display a less favorable lipid profile already shortly postnatal. Maternal low protein diet (LPD) during gestation is a well-defined model of fetal programming in rodents

  9. Increased expression of RXRα in dementia: an early harbinger for the cholesterol dyshomeostasis?

    Directory of Open Access Journals (Sweden)

    Katsel Pavel

    2010-09-01

    Full Text Available Abstract Background Cholesterol content of cerebral membranes is tightly regulated by elaborate mechanisms that balance the level of cholesterol synthesis, uptake and efflux. Among the conventional regulatory elements, a recent research focus has been nuclear receptors, a superfamily of ligand-activated transcription factors providing an indispensable regulatory framework in controlling cholesterol metabolism pathway genes. The mechanism of transcriptional regulation by nuclear receptors such as LXRs involves formation of heterodimers with RXRs. LXR/RXR functions as a sensor of cellular cholesterol concentration and mediates cholesterol efflux by inducing the transcription of key cholesterol shuffling vehicles namely, ATP-binding cassette transporter A1 (ABCA1 and ApoE. Results In the absence of quantitative data from humans, the relevance of expression of nuclear receptors and their involvement in cerebral cholesterol homeostasis has remained elusive. In this work, new evidence is provided from direct analysis of human postmortem brain gene and protein expression suggesting that RXRα, a key regulator of cholesterol metabolism is differentially expressed in individuals with dementia. Importantly, RXRα expression showed strong association with ABCA1 and ApoE gene expression, particularly in AD vulnerable regions. Conclusions These findings suggest that LXR/RXR-induced upregulation of ABCA1 and ApoE levels may be the molecular determinants of cholesterol dyshomeostasis and of the accompanying dementia observed in AD.

  10. Lack of Abcg1 results in decreased plasma HDL cholesterol levels and increased biliary cholesterol secretion in mice fed a high cholesterol diet

    NARCIS (Netherlands)

    Wiersma, Harmen; Nijstad, Niels; de Boer, Jan Freark; Out, Ruud; Hogewerf, Wytse; Van Berkel, Theo J.; Kuipers, Folkert; Tietge, Uwe J. F.

    Objective: The ATP Binding Cassette transporter G1 (ABCG1) has been implicated in cholesterol efflux towards HDL and reverse cholesterol transport (RCT). Biliary cholesterol secretion is considered as an important step in RCT. The aim of the present study was to determine the consequences of Abcg1

  11. From blood to gut: Direct secretion of cholesterol via transintestinal cholesterol efflux

    NARCIS (Netherlands)

    Vrins, Carlos L. J.

    2010-01-01

    The reverse cholesterol transport pathway (RCT) is the focus of many cholesterol lowering therapies By way of this pathway, excess cholesterol is collected from peripheral tissues and delivered back to the liver and gastrointestinal tract for excretion from the body For a long time this removal via

  12. Association of a cholesteryl ester transfer protein variant (rs1800777) with fat mass, HDL cholesterol levels, and metabolic syndrome.

    Science.gov (United States)

    de Luis, Daniel; Izaola, Olatz; Primo, David; Gomez, Emilia; Lopez, Juan Jose; Ortola, Ana; Aller, Rocio

    2018-04-25

    There is little evidence of the association between CETP SNPs and obesity and/or related metabolic parameters. To analyze the association of the polymorphism rs1800777 of the CETP gene with anthropometric parameters, lipid profile, metabolic syndrome and its components, and adipokine levels in obese subjects without type 2 diabetes mellitus or hypertension. A population of 1005 obese subjects was analyzed. Electrical bioimpedance was performed, and blood pressure, presence of metabolic syndrome, dietary intake, physical activity, and biochemical tests were recorded. Nine hundred and sixty eight patients (96.3%) had the GG genotype, 37 patients the GA genotype (3.7%) (no AA genotype was detected). Fat mass (delta: 4.4±1.1kg; p=0.04), waist circumference (delta: 5.6±2.1cm; p=0.02), and waist to hip ratio (delta: 0.04±0.01cm; p=0.01) were higher in A allele carriers than in non-A allele carriers. HDL cholesterol levels were lower in A allele carriers than in non-A allele carriers (delta: 4.2±1.0mg/dL; p=0.04). In the logistic regression analysis, the GA genotype was associated to an increased risk of central obesity (OR 7.55, 95% CI 1.10-55.70, p=0.02) and low HDL cholesterol levels (OR 2.46, 95% CI 1.23-4.91, p=0.014). The CETP variant at position +82 is associated to lower HDL cholesterol levels, increased fat mass, and central obesity in obese subjects. These results may suggest a potential role of this variant gene in pathophysiology of adipose tissue. Copyright © 2018 SEEN y SED. Publicado por Elsevier España, S.L.U. All rights reserved.

  13. Adaptive changes of duodenal iron transport proteins in celiac disease.

    Science.gov (United States)

    Barisani, Donatella; Parafioriti, Antonina; Bardella, Maria Teresa; Zoller, Heinz; Conte, Dario; Armiraglio, Elisabetta; Trovato, Cristina; Koch, Robert O; Weiss, Günter

    2004-05-19

    Iron deficiency is a manifestation of celiac disease (CD) usually attributed to a decreased absorptive surface, although no data on the regulation of iron transport under these conditions are currently available. Our aim was to evaluate divalent metal transporter 1 (DMT1), duodenal cytochrome b (Dcytb), ferroportin 1 (FP1), hephaestin, and transferrin receptor 1 (TfR1) expression, as well as iron regulatory protein (IRP) activity in duodenal biopsies from control, anemic, and CD patients. We studied 10 subjects with dyspepsia, 6 with iron-deficiency anemia, and 25 with CD. mRNA levels were determined by real-time PCR, protein expression by Western blotting or immunohistochemistry, and IRP activity by gel shift assay. Our results showed that DMT1, FP1, hephaestin, and TfR1 mRNA levels were significantly increased in CD patients with reduced body iron stores compared with controls, similar to what was observed in anemic patients. Protein expression paralleled the mRNAs changes. DMT1 protein expression was localized in differentiated enterocytes at the villi tips in controls, whereas with iron deficiency it was observed throughout the villi. FP1 expression was localized on the basolateral membrane of enterocytes and increased with low iron stores. TfR1 was localized in the crypts in controls but also in the villi with iron deficiency. These changes were paralleled by IRP activity, which increased in all iron-deficient subjects. We conclude that duodenal DMT1, FP1, hephaestin, and TfR1 expression and IRP activity, thus the iron absorption capacity, are upregulated in CD patients as a consequence of iron deficiency, whereas the increased enterocyte proliferation observed in CD has no effect on iron uptake regulation.

  14. Changes in plasma low-density lipoprotein (LDL)- and high-density lipoprotein cholesterol in hypo- and hyperthyroid patients are related to changes in free thyroxine, not to polymorphisms in LDL receptor or cholesterol ester transfer protein genes

    NARCIS (Netherlands)

    Diekman, M. J.; Anghelescu, N.; Endert, E.; Bakker, O.; Wiersinga, W. M.

    2000-01-01

    Thyroid function disorders lead to changes in lipoprotein metabolism. Both plasma low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) increase in hypothyroidism and decrease in hyperthyroidism. Changes in LDL-C relate to altered clearance of LDL particles

  15. The cholesterol-lowering effects of oat varieties based on their difference in the composition of proteins and lipids

    OpenAIRE

    Guo, Lina; Tong, Li-Tao; Liu, Liya; Zhong, Kui; Qiu, Ju; Zhou, Sumei

    2014-01-01

    Background The aim of present study is to investigate the hypocholesterolemic effects of the oat components other than the β-glucan in rats fed with a hypercholesterolemic diet. Methods Four-week-old male Wister rats were divided into 6 groups of 7 rats each with similar mean body weights and serum cholesterol concentrations. Rats were fed with the experimental diets containing 10% oats flour for 30 days. Food intake was recorded and monitored everyday to ensure the similar contents of protei...

  16. Multidrug and toxin extrusion proteins mediate cellular transport of cadmium.

    Science.gov (United States)

    Yang, Hong; Guo, Dong; Obianom, Obinna N; Su, Tong; Polli, James E; Shu, Yan

    2017-01-01

    Cadmium (Cd) is an environmentally prevalent toxicant posing increasing risk to human health worldwide. As compared to the extensive research in Cd tissue accumulation, little was known about the elimination of Cd, particularly its toxic form, Cd ion (Cd 2+ ). In this study, we aimed to examine whether Cd 2+ is a substrate of multidrug and toxin extrusion proteins (MATEs) that are important in renal xenobiotic elimination. HEK-293 cells overexpressing the human MATE1 (HEK-hMATE1), human MATE2-K (HEK-hMATE2-K) and mouse Mate1 (HEK-mMate1) were used to study the cellular transport and toxicity of Cd 2+ . The cells overexpressing MATEs showed a 2-4 fold increase of Cd 2+ uptake that could be blocked by the MATE inhibitor cimetidine. A saturable transport profile was observed with the Michaelis-Menten constant (K m ) of 130±15.8μM for HEK-hMATE1; 139±21.3μM for HEK-hMATE2-K; and 88.7±13.5μM for HEK-mMate1, respectively. Cd 2+ could inhibit the uptake of metformin, a substrate of MATE transporters, with the half maximal inhibitory concentration (IC 50 ) of 97.5±6.0μM, 20.2±2.6μM, and 49.9±6.9μM in HEK-hMATE1, HEK-hMATE2-K, and HEK-mMate1 cells, respectively. In addition, hMATE1 could transport preloaded Cd 2+ out of the HEK-hMATE1 cells, thus resulting in a significant decrease of Cd 2+ -induced cytotoxicity. The present study has provided the first evidence supporting that MATEs transport Cd 2+ and may function as cellular elimination machinery in Cd intoxication. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Serotonin transporter (SERT and translocator protein (TSPO expression in the obese ob/ob mouse

    Directory of Open Access Journals (Sweden)

    Santini Ferruccio

    2011-02-01

    Full Text Available Abstract Background An ever growing body of evidences is emerging concerning metabolism hormones, neurotransmitters or stress-related biomarkers as effective modulators of eating behavior and body weight in mammals. The present study sought at examining the density and affinity of two proteins related to neurotransmission and cell metabolism, the serotonin transporter SERT and the cholesterol import-benzodiazepine site TSPO (translocator protein, in a rodent leptin-lacking mutant, the obese ob/ob mouse. Binding studies were thus carried out in brain or peripheral tissues, blood platelets (SERT and kidneys (TSPO, of ob/ob and WT mice supplied with a standard diet, using the selective radiochemical ligands [3H]-paroxetine and [3H]-PK11195. Results We observed comparable SERT number or affinity in brain and platelets of ob/ob and WT mice, whilst a significantly higher [3H]-PK11195 density was reported in the brain of ob/ob animals. TSPO binding parameters were similar in the kidneys of all tested mice. By [3H]-PK11195 autoradiography of coronal hypothalamic-hippocampal sections, an increased TSPO signal was detected in the dentate gyrus (hippocampus and choroids plexus of ob/ob mice, without appreciable changes in the cortex or hypothalamic-thalamic regions. Conclusions These findings show that TSPO expression is up-regulated in cerebral regions of ob/ob leptin-deficient mice, suggesting a role of the translocator protein in leptin-dependent CNS trophism and metabolism. Unchanged SERT in mutant mice is discussed herein in the context of previous literature as the forerunner to a deeper biochemical investigation.

  18. Effect of soy protein on swine intestinal lipoproteins

    International Nuclear Information System (INIS)

    Ho, H.T.

    1987-01-01

    Hypocholesterolemic effect of soy protein appears to be the result of reduced cholesterol absorption and enhanced cholesterol excretion. The objective of this study is to delineate the underlying mechanism of soy protein effect on cholesterol absorption. At the end of a 5-week soy-protein or casein diet, swine were subjected to cannulation of mesenteric lymph duct under halothane anesthesia. A single dose of 250 μCi [ 14 C]-cholesterol and 10 mCi [ 3 H]-leucine was infused into the upper jejunum two hours after one-fifth of daily food was given. Then lymph was collected hourly for three hours and the lipoprotein fractions were separated by ultracentrifugation. SDS-PAGE (5%) was used to measure the concentrations of individual apoproteins by densitometric scanning. The three-hour lymphatic transport of cholesterol in casein-fed swine was significantly higher than in those fed soy protein. Triglyceride transports were similar in two groups. The [ 3 H]-leucine incorporation study revealed that transport of apo B-48 bore a significant positive relationship to transport of cholesterol in both chylomicron and VLDL fractions of mesenteric lymph. A greater apo B-48 secretion with higher specific activity was probably responsible for the greater transport of cholesterol in chylomicrons in casein-fed swine. On the other hand, the lesser cholesterol transport in chylomicrons in soy protein-fed swine was probably caused by lower apo B-48 secretion. Similarly, the transport of lymph VLDL cholesterol in swine fed casein or soy protein paralleled the amount of accompanying apo B-48. Dietary proteins probably influence the intestinal synthesis of apo B-48 which in turn affects cholesterol transport into the lymphatics

  19. Protein-based microhydraulic transport for controllable actuation

    Science.gov (United States)

    Sundaresan, Vishnu Baba; Leo, Donald J.

    2006-03-01

    Plants have the ability to develop large mechanical force from chemical energy available with bio-fuels. The energy released by the cleavage of a terminal phosphate ion during the hydrolysis of a bio- fuel assists the transport of ions and fluids in cellular homeostasis. Materials that develop pressure and hence strain similar to the response of plants to an external stimuli are classified as nastic materials. Calculations for controlled actuation of an active material inspired by biological transport mechanism demonstrated the feasibility of developing such a material with actuation energy densities on the order of 100 kJ/m 3. The mathematical model for a simplified proof of concept actuator referred to as micro hydraulic actuator uses ion transporters extracted from plants reconstituted on a synthetic bilayer lipid membrane (BLM). Thermodynamic model of the concept actuator predicted the ability to develop 5 percent normalized deformation in thickness of the micro- hydraulic actuator. Controlled fluid transport through AtSUT4 (Proton-sucrose co-transporter from Arabidopsis thaliana) reconstituted on a 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-[Phospho-L- Serine] (Sodium Salt) (POPS), 1-Palmitoyl-2-Oleoyl-sn-Glycero-3- Phosphoethanolamine (POPE) BLM on a porous lead silicate glass plate (50μm with 61μm pitch) was driven by proton gradient. Bulk fluid flux of 1.2 μl/min was observed for each microliter of AtSUT4 transporter suspension (16.6 mg/ml in pH7.0 medium) reconstituted on the BLM. The flux rate is observed to be dependent on the concentration of sucrose present in pH4 buffer. Flux rate of 10 μl/min is observed for 5 mM sucrose in the first 10 minutes. The observed flux scales linearly with BLM area and the amount of proteins reconstituted on the lipid membrane. This article details the next step in the development of the micro hydraulic actuator - fluid transport driven by exergonic Adenosine triphosphate (ATP) hydrolysis reaction in the presence of ATP

  20. Egg-Yolk Sphingomyelin and Phosphatidylcholine Attenuate Cholesterol Absorption in Caco-2 Cells.

    Science.gov (United States)

    Yang, Fang; Chen, Guoxun; Ma, Meihu; Qiu, Ning; Zhu, Lingjiao; Li, Jing

    2018-02-01

    Phospholipids have been shown to modulate intestinal cholesterol absorption in cells and animals, a process that is regulated by several transporter proteins. Of these proteins, Niemann-Pick C1-Like 1 (NPC1L1) is a major contributor to this process. The mechanism by which phospholipids modulate cholesterol absorption remains unknown. Here, we evaluate the effects of egg-yolk phospholipids on cholesterol absorption and transport in human colon carcinoma cell line (Caco-2 cells) and on the expression of NPC1L1 and others proteins associated with cholesterol absorption (ABCG5, ABCG8, ABCA1, ACAT2, MTP, CAV-1, ANX-2). The roles of SREBP-1 and SREBP-2 in this process were also investigated. The results show that egg-yolk sphingomyelin (CerPCho) and phosphatidylcholine (PtdCho) inhibit cholesterol transport in the Caco-2 monolayer in a dose-dependent manner. These might be due to the decrease of the cholesterol solubility in micelles as well as to the increases in the micellar sizes and the bile acid-binding capacity. Furthermore, the treatments with egg-yolk CerPCho or PtdCho at 1.2 mmol/L reduced the expression levels of NPC1L1 protein to 21 or 22%, respectively, and its mRNA to 9 or 31% of that in the control group (p egg-yolk PtdCho and CerPCho on the mRNA levels of SREBP-1, and SREBP-2. These results suggest that the inhibitory effect of egg-yolk CerPCho and PtdCho on cholesterol transport might be due to their interference with the physicochemical properties of micelles and their regulations on the expression of the NPC1L1 gene. © 2018 AOCS.

  1. Urea transporter proteins as targets for small-molecule diuretics.

    Science.gov (United States)

    Esteva-Font, Cristina; Anderson, Marc O; Verkman, Alan S

    2015-02-01

    Conventional diuretics such as furosemide and thiazides target salt transporters in kidney tubules, but urea transporters (UTs) have emerged as alternative targets. UTs are a family of transmembrane channels expressed in a variety of mammalian tissues, in particular the kidney. UT knockout mice and humans with UT mutations exhibit reduced maximal urinary osmolality, demonstrating that UTs are necessary for the concentration of urine. Small-molecule screening has identified potent and selective inhibitors of UT-A, the UT protein expressed in renal tubule epithelial cells, and UT-B, the UT protein expressed in vasa recta endothelial cells. Data from UT knockout mice and from rodents administered UT inhibitors support the diuretic action of UT inhibition. The kidney-specific expression of UT-A1, together with high selectivity of the small-molecule inhibitors, means that off-target effects of such small-molecule drugs should be minimal. This Review summarizes the structure, expression and function of UTs, and looks at the evidence supporting the validity of UTs as targets for the development of salt-sparing diuretics with a unique mechanism of action. UT-targeted inhibitors may be useful alone or in combination with conventional diuretics for therapy of various oedemas and hyponatraemias, potentially including those refractory to treatment with current diuretics.

  2. Golgi localized barley MTP8 proteins facilitate Mn transport

    DEFF Research Database (Denmark)

    Pedas, Pai Rosager; Schiller, Michaela; Hegelund, Josefine Nymark

    2014-01-01

    Many metabolic processes in plants are regulated by manganese (Mn) but limited information is available on the molecular mechanisms controlling cellular Mn homeostasis. In this study, a yeast assay was used to isolate and characterize two genes, MTP8.1 and MTP8.2 , which encode membrane-bound pro......Many metabolic processes in plants are regulated by manganese (Mn) but limited information is available on the molecular mechanisms controlling cellular Mn homeostasis. In this study, a yeast assay was used to isolate and characterize two genes, MTP8.1 and MTP8.2 , which encode membrane...... in yeast, MTP8.1 and MTP8.2 were found to be Mn transporters catalysing Mn efflux in a similar manner as the Golgi localized endogenous yeast protein Pmr1p. The level of MTP8.1 transcripts in barley roots increased with external Mn supply ranging from deficiency to toxicity, while MTP8.2 transcripts......8 proteins are involved in Mn loading to the Golgi apparatus and play a role in Mn homeostasis by delivering Mn to Mn-dependent enzymes and/or by facilitating Mn efflux via secretory vesicles. This study highlights the importance of MTP transporters in Mn homeostasis and is the first report of Golgi...

  3. [Glucose transporter protein type 1 (GLUT-1) deficiency syndrome].

    Science.gov (United States)

    Ramm-Pettersen, Anette; Selmer, Kaja Kristine; Nakken, Karl O

    2011-05-06

    Glucose is the brain's main source of energy. To pass the blood-brain barrier, glucose transporter protein type 1 (GLUT-1) is essential. Mutations in the SLC2A1 gene which codes for GLUT-1 may therefore compromise the supply of glucose to the brain. The aim of this review is to describe the clinical consequences of such mutations, with special emphasis on GLUT-1 encephalopathy. This review is based on a non-systematic literature search in PubMed and the authors' experience within the field. Epileptic or epilepsy-like are usually the first symptom in children with the GLUT-1 deficiency syndrome. Later on these children suffer delayed psychomotor development, microcephaly, ataxia, spasticity or movement disorders. EEG abnormalities may develop. GLUT-1 deficiency syndrome should be suspected in children with epilepsy-like seizures and delayed development combined with a low content of glucose in spinal fluid. The diagnosis is confirmed by genetic testing. Treatment is a ketogenic diet, as ketone bodies pass the blood-brain barrier using other transport proteins than GLUT-1. GLUT-1-deficiency syndrome is a rare metabolic encephalopathy which is not well known and probably underdiagnosed. An early diagnosis and early start of a ketogenic diet may give these children a normal or nearly normal life.

  4. Export of recombinant proteins in Escherichia coli using ABC transporter with an attached lipase ABC transporter recognition domain (LARD

    Directory of Open Access Journals (Sweden)

    Moon Yuseok

    2009-01-01

    Full Text Available Abstract Background ATP binding cassette (ABC transporter secretes the protein through inner and outer membranes simultaneously in gram negative bacteria. Thermostable lipase (TliA of Pseudomonas fluorescens SIK W1 is secreted through the ABC transporter. TliA has four glycine-rich repeats (GGXGXD in its C-terminus, which appear in many ABC transporter-secreted proteins. From a homology model of TliA derived from the structure of P. aeruginosa alkaline protease (AprA, lipase ABC transporter domains (LARDs were designed for the secretion of fusion proteins. Results The LARDs included four glycine-rich repeats comprising a β-roll structure, and were added to the C-terminus of test proteins. Either Pro-Gly linker or Factor Xa site was added between fusion proteins and LARDs. We attached different length of LARDs such as LARD0, LARD1 or whole TliA (the longest LARD to three types of proteins; green fluorescent protein (GFP, epidermal growth factor (EGF and cytoplasmic transduction peptide (CTP. These fusion proteins were expressed in Escherichia coli together with ABC transporter of either P. fluorescens or Erwinia chrysanthemi. Export of fusion proteins with the whole TliA through the ABC transporter was evident on the basis of lipase enzymatic activity. Upon supplementation of E. coli with ABC transporter, GFP-LARDs and EGF-LARDs were excreted into the culture supernatant. Conclusion The LARDs or whole TliA were attached to C-termini of model proteins and enabled the export of the model proteins such as GFP and EGF in E. coli supplemented with ABC transporter. These results open the possibility for the extracellular production of recombinant proteins in Pseudomonas using LARDs or TliA as a C-terminal signal sequence.

  5. Cellular Cholesterol Facilitates the Postentry Replication Cycle of Herpes Simplex Virus 1.

    Science.gov (United States)

    Wudiri, George A; Nicola, Anthony V

    2017-07-15

    initial entry and transport of viral capsids to the nucleus. Viral protein expression, encapsidation of the viral genome, and the release of mature virions were impacted by the reduction of cellular cholesterol. Cholesterol was also critical for cell-to-cell spread of infection. These findings provide new insights into the cholesterol dependence of HSV-1 replication. Copyright © 2017 American Society for Microbiology.

  6. Nanoscale Membrane Domain Formation Driven by Cholesterol

    DEFF Research Database (Denmark)

    Javanainen, Matti; Martinez-Seara, Hector; Vattulainen, Ilpo

    2017-01-01

    Biological membranes generate specific functions through compartmentalized regions such as cholesterol-enriched membrane nanodomains that host selected proteins. Despite the biological significance of nanodomains, details on their structure remain elusive. They cannot be observed via microscopic...... dipalmitoylphosphatidylcholine and cholesterol - the "minimal standard" for nanodomain formation. The simulations reveal how cholesterol drives the formation of fluid cholesterol-rich nanodomains hosting hexagonally packed cholesterol-poor lipid nanoclusters, both of which show registration between the membrane leaflets....... The complex nanodomain substructure forms when cholesterol positions itself in the domain boundary region. Here cholesterol can also readily flip-flop across the membrane. Most importantly, replacing cholesterol with a sterol characterized by a less asymmetric ring region impairs the emergence of nanodomains...

  7. Lipid Absorption Defects in Intestine-specific Microsomal Triglyceride Transfer Protein and ATP-binding Cassette Transporter A1-deficient Mice*

    Science.gov (United States)

    Iqbal, Jahangir; Parks, John S.; Hussain, M. Mahmood

    2013-01-01

    We have previously described apolipoprotein B (apoB)-dependent and -independent cholesterol absorption pathways and the role of microsomal triglyceride transfer protein (MTP) and ATP-binding cassette transporter A1 (ABCA1) in these pathways. To assess the contribution of these pathways to cholesterol absorption and to determine whether there are other pathways, we generated mice that lack MTP and ABCA1, individually and in combination, in the intestine. Intestinal deletions of Mttp and Abca1 decreased plasma cholesterol concentrations by 45 and 24%, respectively, whereas their combined deletion reduced it by 59%. Acute cholesterol absorption was reduced by 28% in the absence of ABCA1, and it was reduced by 92–95% when MTP was deleted in the intestine alone or together with ABCA1. MTP deficiency significantly reduced triglyceride absorption, although ABCA1 deficiency had no effect. ABCA1 deficiency did not affect cellular lipids, but Mttp deficiency significantly increased intestinal levels of triglycerides and free fatty acids. Accumulation of intestinal free fatty acids, but not triglycerides, in Mttp-deficient intestines was prevented when mice were also deficient in intestinal ABCA1. Combined deficiency of these genes increased intestinal fatty acid oxidation as a consequence of increased expression of peroxisome proliferator-activated receptor-γ (PPARγ) and carnitine palmitoyltransferase 1α (CPT1α). These studies show that intestinal MTP and ABCA1 are critical for lipid absorption and are the main determinants of plasma and intestinal lipid levels. Reducing their activities might lower plasma lipid concentrations. PMID:24019513

  8. Total Cholesterol and Cholesterol Species Determination

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Wei Zou ### Abstract Total cholesterol and cholesterol species analysis are critical in cardiovascular disease research. The protocol shows procedures that can be used for analysing tissue lipid extracts, lymph, bile or serum. ### Reagents 1. Free cholesterol standard solution (1 mg/mL in ethanol) - Cholesterol palmitate standard solution (1 mg/mL in chloroform): Add 106.383 mg of cholesterol palmitate (94%, Sigma) into a volumetric flask, top with chloroform. ...

  9. Regulation of dopamine transporter function by protein-protein interactions: new discoveries and methodological challenges

    DEFF Research Database (Denmark)

    Eriksen, Jacob; Jørgensen, Trine Nygaard; Gether, Ulrik

    2010-01-01

    The dopamine transporter (DAT) plays a key role in regulating dopaminergic signalling in the brain by mediating rapid clearance of dopamine from the synaptic clefts. The psychostimulatory actions of cocaine and amphetamine are primarily the result of a direct interaction of these compounds with DAT...... cells have also recently become available such as fluorescently tagged cocaine analogues and fluorescent substrates. Here we review the current knowledge about the role of protein-protein interactions in DAT regulation as well as we describe the most recent methodological developments that have been...

  10. Whole Soy Flour Incorporated into a Muffin and Consumed at 2 Doses of Soy Protein Does Not Lower LDL Cholesterol in a Randomized, Double-Blind Controlled Trial of Hypercholesterolemic Adults.

    Science.gov (United States)

    Padhi, Emily Mt; Blewett, Heather J; Duncan, Alison M; Guzman, Randolph P; Hawke, Aileen; Seetharaman, Koushik; Tsao, Rong; Wolever, Thomas Ms; Ramdath, D Dan

    2015-12-01

    Soy protein may reduce coronary heart disease (CHD) risk by lowering LDL cholesterol, but few studies have assessed whether whole soy flour displays a similar effect. The aim of this study was to assess the dose effect of whole soy flour incorporated into muffins on plasma LDL cholesterol in hypercholesterolemic adults. Adults aged 30-70 y (n = 243) with elevated LDL cholesterol (≥3.0 and ≤5.0 mmol/L) were stratified by LDL cholesterol and randomly assigned to consume 2 soy muffins containing 25 g soy protein [high-dose soy (HDS)], 1 soy and 1 wheat muffin containing 12.5 g soy protein and 12.5 g whey protein [low-dose soy (LDS)], or 2 wheat muffins containing 25 g whey protein (control) daily for 6 wk while consuming a self-selected diet. Fasting blood samples were collected at weeks 0, 3, and 6 for analysis of plasma lipids [total, LDL, and HDL cholesterol and triglycerides (TGs)], glucose, insulin, C-reactive protein (CRP), and isoflavones. Blood pressures also were measured. Dietary intake was assessed at weeks 0 and 4 with the use of 3 d food records. Treatment effects were assessed with the use of intention-to-treat analysis with multiple imputation and LDL cholesterol as the primary outcome. In total, 213 (87.6%) participants completed the trial. Participants were primarily Caucasian (83%) and mostly female (63%), with a mean ± SD body mass index (in kg/m2) of 28.0 ± 4.6 and systolic and diastolic blood pressures of 122 ± 16 and 77 ± 11 mm Hg, respectively. Despite a dose-dependent increase in plasma isoflavones (P LDL cholesterol compared with control (mean ± SEM changes: control, -0.04 ± 0.05 mmol/L; HDS, 0.01 ± 0.05 mmol/L; and LDS, -0.04 ± 0.06 mmol/L). There were no significant treatment effects on total or HDL cholesterol, TGs, CRP, homeostatic model assessment of insulin resistance, blood pressure, or the Framingham 10-y CHD risk score. Consuming 12.5 or 25 g protein from defatted soy flour incorporated into muffins does not reduce LDL

  11. Whole Soy Flour Incorporated into a Muffin and Consumed at 2 Doses of Soy Protein Does Not Lower LDL Cholesterol in a Randomized, Double-Blind Controlled Trial of Hypercholesterolemic Adults12

    Science.gov (United States)

    Padhi, Emily MT; Blewett, Heather J; Duncan, Alison M; Guzman, Randolph P; Hawke, Aileen; Seetharaman, Koushik; Tsao, Rong; Wolever, Thomas MS; Ramdath, D Dan

    2015-01-01

    Background: Soy protein may reduce coronary heart disease (CHD) risk by lowering LDL cholesterol, but few studies have assessed whether whole soy flour displays a similar effect. Objective: The aim of this study was to assess the dose effect of whole soy flour incorporated into muffins on plasma LDL cholesterol in hypercholesterolemic adults. Methods: Adults aged 30–70 y (n = 243) with elevated LDL cholesterol (≥3.0 and ≤5.0 mmol/L) were stratified by LDL cholesterol and randomly assigned to consume 2 soy muffins containing 25 g soy protein [high-dose soy (HDS)], 1 soy and 1 wheat muffin containing 12.5 g soy protein and 12.5 g whey protein [low-dose soy (LDS)], or 2 wheat muffins containing 25 g whey protein (control) daily for 6 wk while consuming a self-selected diet. Fasting blood samples were collected at weeks 0, 3, and 6 for analysis of plasma lipids [total, LDL, and HDL cholesterol and triglycerides (TGs)], glucose, insulin, C-reactive protein (CRP), and isoflavones. Blood pressures also were measured. Dietary intake was assessed at weeks 0 and 4 with the use of 3 d food records. Treatment effects were assessed with the use of intention-to-treat analysis with multiple imputation and LDL cholesterol as the primary outcome. Results: In total, 213 (87.6%) participants completed the trial. Participants were primarily Caucasian (83%) and mostly female (63%), with a mean ± SD body mass index (in kg/m2) of 28.0 ± 4.6 and systolic and diastolic blood pressures of 122 ± 16 and 77 ± 11 mm Hg, respectively. Despite a dose-dependent increase in plasma isoflavones (P < 0.001), neither HDS nor LDS had a significant effect on LDL cholesterol compared with control (mean ± SEM changes: control, −0.04 ± 0.05 mmol/L; HDS, 0.01 ± 0.05 mmol/L; and LDS, −0.04 ± 0.06 mmol/L). There were no significant treatment effects on total or HDL cholesterol, TGs, CRP, homeostatic model assessment of insulin resistance, blood pressure, or the Framingham 10-y CHD risk score

  12. Effect of P/S ratio (0.5 vs 0.9) on hepatic LDL transport at three levels of dietary cholesterol in cynomolgus macaques

    International Nuclear Information System (INIS)

    Hunt, C.E.; Funk, G.M.; Turley, S.D.; Spady, D.K.; Dietschy, J.M.

    1990-01-01

    Interaction between dietary polyunsaturated to saturated (P/S) fatty acid ratio and cholesterol (C) was studied in 6 groups of male cynomolgus macaques fed diets (oleic acid constant) for 72 weeks as follows (C mg/Cal-P/S): (1), 0.06 - 0.5; (2), 0.06-0.9, (3), 0.28-0.5; (4), 0.28-0.9; (5), 2,35-0.5; (6), 2,35-0.9. Plasma C was proportional to dietary C and was affected significantly by P/S in 1 and 2 only. Mean plasma C (mg/dl) at 72 weeks was: (1) 158; (2) 117; (3) 320; (4) 284; (5) 602; (6) 601. LDL-C was significantly higher in (1) than in (2) (90 vs 65 mg/dl). In vivo LDL turnover studies showed that LDL clearance was suppressed by excess dietary C and by saturated fats in low C diets. Receptor-independent clearance was relatively constant. Hepatic LDL transport was determined after injection of 125I-cellobiose-LDL. Hepatic LDL-C uptake was greater in (2) than in (1). LDL-C synthesis was reduced in (4) and (6) compared to (3) and (5), respectively. The authors conclude that (i) hepatic LDL receptor activity is altered by degree of saturation in dietary triglycerides when dietary C is minimal and (ii) saturated triglycerides enhance LDL-C synthesis when dietary C is ample in this model

  13. Cholesterol worships a new idol.

    Science.gov (United States)

    Schulman, Ira G

    2009-12-01

    The growing worldwide epidemic of cardiovascular disease suggests that new therapeutic strategies are needed to complement statins in the lowering of cholesterol levels. In a recent paper in Science, Tontonoz and colleagues have identified Idol as a protein that can control cholesterol levels by regulating the stability of the low-density lipoprotein receptor; inhibiting the activity of Idol could provide novel approaches for the treatment of cardiovascular disease.

  14. Interaction of cholesterol ester transfer protein polymo- rphisms, body mass index, and birth weight with the risk of dyslipidemia in children and adolescents: the CASPIAN-III study

    Directory of Open Access Journals (Sweden)

    Motahar Heidari-Beni

    2015-11-01

    Full Text Available Objective(s: This study aims to investigate joint association between cholesterol ester transfer protein (CETP polymorphisms and body mass index (BMI or birth weight with the risk of dyslipidemia in Iranian children and adolescents. Materials and Methods:This study was conducted as a sub-study of the “school-based nationwide health survey” (CASPIAN-III. We randomly selected 750 samples from the whole blood samples. Real-time PCR and high resolution melt (HRM analysis were performed to determine Taq1B (rs708272 and A373P (rs5880 polymorphisms. Results:Taq1B polymorphism increased HDL-C, and total cholesterol (TC as well as decreased triglyceride and LDL-C concentrations. LDL-C and triglyceride levels were significantly higher and HDL-C and TC levels were significantly lower among those with A373P polymorphism. CT/TT genotype in Taq1B polymorphism showed a protective effect on dyslipidemia (OR= 0.12, 95%CI: 0.07-0.20. G allele of A373P polymorphism increased the risk of dyslipidemia (OR=4.10, 95%CI: 2.14, 7.83 after adjusting the confounders. We observed interactive effects of CETP gene polymorphisms and BMI or birth weight on dyslipidemia. Conclusion:Findings showed Taq1B polymorphism might have a protective effect and A373P polymorphism had deleterious effect on dyslipidemia in Iranian children and adolescents. These associations interacted with BMI and birth weight.

  15. Dietary Wheat Bran Oil Is Equally as Effective as Rice Bran Oil in Reducing Plasma Cholesterol.

    Science.gov (United States)

    Lei, Lin; Chen, Jingnan; Liu, Yuwei; Wang, Lijun; Zhao, Guohua; Chen, Zhen-Yu

    2018-03-21

    Rice bran oil (RBO) possesses a plasma cholesterol-lowering activity, while effect of wheat bran oil (WBO) on plasma cholesterol remains unknown. The present study compared the cholesterol-lowering activity of WBO with that of RBO in hamsters. Fifty-four male hamsters were divided into seven groups fed either a noncholesterol diet (NCD) or one of six high-cholesterol diets, namely HCD diet (0.2% cholesterol +9.5% lard), HCD+C diet (0.2% cholesterol +9.5% lard +0.5% cholestyramine), WL diet (0.2% cholesterol +4.8% Lard +4.8% WBO), WH diet (0.2% cholesterol +9.5% WBO), RL diet (0.2% cholesterol +4.8% Lard +4.8% RBO), and RH diet (0.2% cholesterol +9.5% RBO). Plasma total cholesterol (TC) in HCD group was 327.4 ± 31.8 mg/dL, while plasma TC in two WBO and two RBO groups was 242.2 ± 20.8, 243.1 ± 31.7, 257.1 ± 16.3, and 243.4 ± 46.0 mg/dL, respectively, leading to a decrease in plasma TC by 22-26% ( P cholesterol-lowering potency was seen between WBO and RBO. Plasma cholesterol-lowering activity of WBO and RBO was accompanied by down-regulation of hepatic 3-hydroxy-3-methylglutaryl-CoA reductase and fatty acid synthase, while up-regulation of cholesterol-7α-hydroxylase. WL, WH, RL, and RH diets increased the fecal excretion of total neutral sterols by 72.8%, 106.9%, 5.4%, and 36.8% ( P cholesterol absorption via down-regulation of intestinal Niemann-Pick C1 like 1 protein, acyl CoA:cholesterol acyltransferase 2, and ATP binding cassette transporter 5. In summary, WBO was equally effective as RBO in decreasing plasma cholesterol in hypercholesterolemia hamsters.

  16. Oxidized Low-density Lipoprotein (ox-LDL) Cholesterol Induces the Expression of miRNA-223 and L-type Calcium Channel Protein in Atrial Fibrillation

    Science.gov (United States)

    He, Fengping; Xu, Xin; Yuan, Shuguo; Tan, Liangqiu; Gao, Lingjun; Ma, Shaochun; Zhang, Shebin; Ma, Zhanzhong; Jiang, Wei; Liu, Fenglian; Chen, Baofeng; Zhang, Beibei; Pang, Jungang; Huang, Xiuyan; Weng, Jiaqiang

    2016-08-01

    Atrial fibrillation (AF) is the most common sustained arrhythmia causing high morbidity and mortality. While changing of the cellular calcium homeostasis plays a critical role in AF, the L-type calcium channel α1c protein has suggested as an important regulator of reentrant spiral dynamics and is a major component of AF-related electrical remodeling. Our computational modeling predicted that miRNA-223 may regulate the CACNA1C gene which encodes the cardiac L-type calcium channel α1c subunit. We found that oxidized low-density lipoprotein (ox-LDL) cholesterol significantly up-regulates both the expression of miRNA-223 and L-type calcium channel protein. In contrast, knockdown of miRNA-223 reduced L-type calcium channel protein expression, while genetic knockdown of endogenous miRNA-223 dampened AF vulnerability. Transfection of miRNA-223 by adenovirus-mediated expression enhanced L-type calcium currents and promoted AF in mice while co-injection of a CACNA1C-specific miR-mimic counteracted the effect. Taken together, ox-LDL, as a known factor in AF-associated remodeling, positively regulates miRNA-223 transcription and L-type calcium channel protein expression. Our results implicate a new molecular mechanism for AF in which miRNA-223 can be used as an biomarker of AF rheumatic heart disease.

  17. Scavenger Receptor Class B Type I Mediates Biliary Cholesterol Secretion Independent of ATP-Binding Cassette Transporter g5/g8 in Mice

    NARCIS (Netherlands)

    Wiersma, Harmen; Gatti, Alberto; Nijstad, Niels; Elferink, Ronald P. J. Oude; Kuipers, Folkert; Tietge, Uwe J. F.

    2009-01-01

    Scavenger receptor class B type I (SR-BI) mediates selective uptake of cholesterol from high-density lipoprotein (HDL) particles by the liver and influences biliary cholesterol secretion. However, it is not dear, if this effect is direct or indirect. The aim of this study was to determine the impact

  18. The twin arginine protein transport pathway exports multiple virulence proteins in the plant pathogen Streptomyces scabies.

    Science.gov (United States)

    Joshi, Madhumita V; Mann, Stefan G; Antelmann, Haike; Widdick, David A; Fyans, Joanna K; Chandra, Govind; Hutchings, Matthew I; Toth, Ian; Hecker, Michael; Loria, Rosemary; Palmer, Tracy

    2010-07-01

    Summary Streptomyces scabies is one of a group of organisms that causes the economically important disease potato scab. Analysis of the S. scabies genome sequence indicates that it is likely to secrete many proteins via the twin arginine protein transport (Tat) pathway, including several proteins whose coding sequences may have been acquired through horizontal gene transfer and share a common ancestor with proteins in other plant pathogens. Inactivation of the S. scabies Tat pathway resulted in pleiotropic phenotypes including slower growth rate and increased permeability of the cell envelope. Comparison of the extracellular proteome of the wild type and DeltatatC strains identified 73 predicted secretory proteins that were present in reduced amounts in the tatC mutant strain, and 47 Tat substrates were verified using a Tat reporter assay. The DeltatatC strain was almost completely avirulent on Arabidopsis seedlings and was delayed in attaching to the root tip relative to the wild-type strain. Genes encoding 14 candidate Tat substrates were individually inactivated, and seven of these mutants were reduced in virulence compared with the wild-type strain. We conclude that the Tat pathway secretes multiple proteins that are required for full virulence.

  19. Alcohol consumption, TaqIB polymorphism of cholesteryl ester transfer protein, high-density lipoprotein cholesterol, and risk of coronary heart disease in men and women

    DEFF Research Database (Denmark)

    Jensen, Majken K; Mukamal, Kenneth J; Overvad, Kim

    2008-01-01

    AIMS: To investigate whether a common polymorphism in the cholesteryl ester transfer protein (CETP) gene modifies the relationship of alcohol intake with high-density lipoprotein cholesterol (HDL-C) and risk of coronary heart disease (CHD). METHODS AND RESULTS: Parallel nested case-control studies...... among women [Nurses' Health Study (NHS)] and men [Health Professionals Follow-up Study (HPFS)] where 246 women and 259 men who developed incident CHD were matched to controls (1:2) on age and smoking. The TaqIB variant and alcohol consumption were associated with higher HDL-C, with the most pronounced...... effects of alcohol among B2 carriers. In the NHS we did not find an inverse association between alcohol and CHD in B2 non-carriers (P trend: 0.5), but did among B2 carriers (P trend

  20. Regional distribution of serotonin transporter protein in postmortem human brain

    International Nuclear Information System (INIS)

    Kish, Stephen J.; Furukawa, Yoshiaki; Chang Lijan; Tong Junchao; Ginovart, Nathalie; Wilson, Alan; Houle, Sylvain; Meyer, Jeffrey H.

    2005-01-01

    Introduction: The primary approach in assessing the status of brain serotonin neurons in human conditions such as major depression and exposure to the illicit drug ecstasy has been the use of neuroimaging procedures involving radiotracers that bind to the serotonin transporter (SERT). However, there has been no consistency in the selection of a 'SERT-free' reference region for the estimation of free and nonspecific binding, as occipital cortex, cerebellum and white matter have all been employed. Objective and Methods: To identify areas of human brain that might have very low SERT levels, we measured, by a semiquantitative Western blotting procedure, SERT protein immunoreactivity throughout the postmortem brain of seven normal adult subjects. Results: Serotonin transporter could be quantitated in all examined brain areas. However, the SERT concentration in cerebellar cortex and white matter were only at trace values, being approximately 20% of average cerebral cortex and 5% of average striatum values. Conclusion: Although none of the examined brain areas are completely free of SERT, human cerebellar cortex has low SERT binding as compared to other examined brain regions, with the exception of white matter. Since the cerebellar cortical SERT binding is not zero, this region will not be a suitable reference region for SERT radioligands with very low free and nonspecific binding. For SERT radioligands with reasonably high free and nonspecific binding, the cerebellar cortex should be a useful reference region, provided other necessary radioligand assumptions are met

  1. A finite element model for protein transport in vivo

    Directory of Open Access Journals (Sweden)

    Montas Hubert J

    2007-06-01

    Full Text Available Abstract Background Biological mass transport processes determine the behavior and function of cells, regulate interactions between synthetic agents and recipient targets, and are key elements in the design and use of biosensors. Accurately predicting the outcomes of such processes is crucial to both enhancing our understanding of how these systems function, enabling the design of effective strategies to control their function, and verifying that engineered solutions perform according to plan. Methods A Galerkin-based finite element model was developed and implemented to solve a system of two coupled partial differential equations governing biomolecule transport and reaction in live cells. The simulator was coupled, in the framework of an inverse modeling strategy, with an optimization algorithm and an experimental time series, obtained by the Fluorescence Recovery after Photobleaching (FRAP technique, to estimate biomolecule mass transport and reaction rate parameters. In the inverse algorithm, an adaptive method was implemented to calculate sensitivity matrix. A multi-criteria termination rule was developed to stop the inverse code at the solution. The applicability of the model was illustrated by simulating the mobility and binding of GFP-tagged glucocorticoid receptor in the nucleoplasm of mouse adenocarcinoma. Results The numerical simulator shows excellent agreement with the analytic solutions and experimental FRAP data. Detailed residual analysis indicates that residuals have zero mean and constant variance and are normally distributed and uncorrelated. Therefore, the necessary and sufficient criteria for least square parameter optimization, which was used in this study, were met. Conclusion The developed strategy is an efficient approach to extract as much physiochemical information from the FRAP protocol as possible. Well-posedness analysis of the inverse problem, however, indicates that the FRAP protocol provides insufficient

  2. Artificial membranes with selective nanochannels for protein transport

    KAUST Repository

    Sutisna, B.

    2016-09-05

    A poly(styrene-b-tert-butoxystyrene-b-styrene) copolymer was synthesized by anionic polymerization and hydrolyzed to poly(styrene-b-4-hydroxystyrene-b-styrene). Lamellar morphology was confirmed in the bulk after annealing. Membranes were fabricated by self-assembly of the hydrolyzed copolymer in solution, followed by water induced phase separation. A high density of pores of 4 to 5 nm diameter led to a water permeance of 40 L m−2 h−1 bar−1 and molecular weight cut-off around 8 kg mol−1. The morphology was controlled by tuning the polymer concentration, evaporation time, and the addition of imidazole and pyridine to stabilize the terpolymer micelles in the casting solution via hydrogen bond complexes. Transmission electron microscopy of the membrane cross-sections confirmed the formation of channels with hydroxyl groups beneficial for hydrogen-bond forming sites. The morphology evolution was investigated by time-resolved grazing incidence small angle X-ray scattering experiments. The membrane channels reject polyethylene glycol with a molecular size of 10 kg mol−1, but are permeable to proteins, such as lysozyme (14.3 kg mol−1) and cytochrome c (12.4 kg mol−1), due to the right balance of hydrogen bond interactions along the channels, electrostatic attraction, as well as the right pore sizes. Our results demonstrate that artificial channels can be designed for protein transport via block copolymer self-assembly using classical methods of membrane preparation.

  3. The dynamin chemical inhibitor dynasore impairs cholesterol trafficking and sterol-sensitive genes transcription in human HeLa cells and macrophages.

    Directory of Open Access Journals (Sweden)

    Emmanuelle Girard

    Full Text Available Intracellular transport of cholesterol contributes to the regulation of cellular cholesterol homeostasis by mechanisms that are yet poorly defined. In this study, we characterized the impact of dynasore, a recently described drug that specifically inhibits the enzymatic activity of dynamin, a GTPase regulating receptor endocytosis and cholesterol trafficking. Dynasore strongly inhibited the uptake of low-density lipoprotein (LDL in HeLa cells, and to a lower extent in human macrophages. In both cell types, dynasore treatment led to the abnormal accumulation of LDL and free cholesterol (FC within the endolysosomal network. The measure of cholesterol esters (CE further showed that the delivery of regulatory cholesterol to the endoplasmic reticulum (ER was deficient. This resulted in the inhibition of the transcriptional control of the three major sterol-sensitive genes, sterol-regulatory element binding protein 2 (SREBP-2, 3-hydroxy-3-methyl-coenzymeA reductase (HMGCoAR, and low-density lipoprotein receptor (LDLR. The sequestration of cholesterol in the endolysosomal compartment impaired both the active and passive cholesterol efflux in HMDM. Our data further illustrate the importance of membrane trafficking in cholesterol homeostasis and validate dynasore as a new pharmacological tool to study the intracellular transport of cholesterol.

  4. Cholesterol Facts and Statistics

    Science.gov (United States)

    ... Blood Pressure Salt Million Hearts® WISEWOMAN Program High Cholesterol Facts Recommend on Facebook Tweet Share Compartir Find ... about high cholesterol in the United States. High Cholesterol in the United States In 2011–2012, 78 ...

  5. Common Misconceptions about Cholesterol

    Science.gov (United States)

    ... Venous Thromboembolism Aortic Aneurysm More Common Misconceptions about Cholesterol Updated:Jan 29,2018 How much do you ... are some common misconceptions — and the truth. High cholesterol isn’t a concern for children. High cholesterol ...

  6. Cholesterol testing and results

    Science.gov (United States)

    Cholesterol test results; LDL test results; VLDL test results; HDL test results; Coronary risk profile results; Hyperlipidemia-results; Lipid disorder test results; Heart disease - cholesterol results

  7. Ezetimibe: Its Novel Effects on the Prevention and the Treatment of Cholesterol Gallstones and Nonalcoholic Fatty Liver Disease

    Directory of Open Access Journals (Sweden)

    Ornella de Bari

    2012-01-01

    Full Text Available The cholesterol absorption inhibitor ezetimibe can significantly reduce plasma cholesterol concentrations by inhibiting the Niemann-Pick C1-like 1 protein (NPC1L1, an intestinal sterol influx transporter that can actively facilitate the uptake of cholesterol for intestinal absorption. Unexpectedly, ezetimibe treatment also induces a complete resistance to cholesterol gallstone formation and nonalcoholic fatty liver disease (NAFLD in addition to preventing hypercholesterolemia in mice on a Western diet. Because chylomicrons are the vehicles with which the enterocytes transport cholesterol and fatty acids into the body, ezetimibe could prevent these two most prevalent hepatobiliary diseases possibly through the regulation of chylomicron-derived cholesterol and fatty acid metabolism in the liver. It is highly likely that there is an intestinal and hepatic cross-talk through the chylomicron pathway. Therefore, understanding the molecular mechanisms whereby cholesterol and fatty acids are absorbed from the intestine could offer an efficacious novel approach to the prevention and the treatment of cholesterol gallstones and NAFLD.

  8. ¹H, ¹³C, and ¹⁵N backbone resonance assignments of the L124D mutant of StAR-related lipid transfer domain protein 4 (StARD4).

    Science.gov (United States)

    Dikiy, Igor; Ramlall, Trudy F; Eliezer, David

    2013-10-01

    Protein-mediated cholesterol trafficking is central to maintaining cholesterol homeostasis in cells. START (Steroidogenic acute regulatory protein-related lipid transfer) domains constitute a sterol and lipid binding motif and the START domain protein StARD4 typifies a small family of mammalian sterol transport proteins. StARD4 consists of a single START domain and has been reported to act as a general cholesterol transporter in cells. However, the structural basis of cholesterol uptake and transport is not well understood and no cholesterol-bound START domain structures have been reported. We have undertaken the study of cholesterol binding and transport by StARD4 using solution state NMR spectroscopy. To this end, we report nearly complete (1)H, (15)N, and (13)C backbone resonance assignments of an inactive but well behaved mutant (L124D) of StARD4.

  9. 1H, 13C, and 15N backbone resonance assignments of the L124D mutant of StAR-related lipid transfer domain protein 4 (StARD4)

    Science.gov (United States)

    Dikiy, Igor; Ramlall, Trudy F.; Eliezer, David

    2012-01-01

    Protein-mediated cholesterol trafficking is central to maintaining cholesterol homeostasis in cells. START (Steroidogenic acute regulatory protein-related lipid transfer) domains constitute a sterol and lipid binding motif and the START domain protein StARD4 typifies a small family of mammalian sterol transport proteins. StARD4 consists of a single START domain and has been reported to act as a general cholesterol transporter in cells. However, the structural basis of cholesterol uptake and transport is not well understood and no cholesterol-bound START domain structures have been reported. We have undertaken the study of cholesterol binding and transport by StARD4 using solution state NMR spectroscopy. To this end, we report nearly complete 1H, 15N, and 13C backbone resonance assignments of an inactive but well behaved mutant (L124D) of StARD4. PMID:22918595

  10. Influence of human diets containing casein, soy protein isolate, and soy protein concentrate on serum cholesterol and lipoproteins in humans, rabbits and rats

    NARCIS (Netherlands)

    Raaij, van J.M.A.

    1982-01-01

    It is well known that feeding animals such as rabbits with semipurified diets containing animal proteins, as for example casein, results in hypercholesterolemia and atherosclerosis. On the other hand, diets containing vegetable proteins such as soybean protein maintain low levels of serum

  11. Dietary cholesterol and the plasma lipids and lipoproteins in the Tarahumara Indians: a people habituated to a low cholesterol diet after weaning.

    Science.gov (United States)

    McMurry, M P; Connor, W E; Cerqueira, M T

    1982-04-01

    Eight Tarahumara Indian men participated in a metabolic study to measure the responsiveness of their plasma cholesterol levels to dietary cholesterol. They were fed isocaloric cholesterol-free and high cholesterol diets containing 20% fat, 15% protein, and 65% carbohydrate calories. On admission to the study, the Tarahumaras had a low mean plasma cholesterol concentration (120 mg/dl), reflecting their habitual low cholesterol diet. After 3 wk of a cholesterol-free diet their cholesterol levels were 113 mg/dl. The men were then fed a high cholesterol diet (1000 mg/day) which increased the mean total plasma cholesterol to 147 mg/dl (p less than 0.01) and also increased the low-density lipoprotein cholesterol concentration. Tarahumaras, habituated to a low cholesterol diet after weaning, had the typical hypercholesterolemic response to a high cholesterol diet that has been previously observed in subjects whose lifelong diet was high in cholesterol content.

  12. Orlistat limits cholesterol intestinal absorption by Niemann-pick C1-like 1 (NPC1L1) inhibition.

    Science.gov (United States)

    Alqahtani, Saeed; Qosa, Hisham; Primeaux, Brian; Kaddoumi, Amal

    2015-09-05

    The known mechanism by which orlistat decreases the absorption of dietary cholesterol is by inhibition of intestinal lipases. The aim of this study was to investigate the ability of orlistat to limit cholesterol absorption by inhibition of the cholesterol transport protein Niemann-Pick C1-like 1 (NPC1L1) as another mechanism of action. In situ rat intestinal perfusion studies were conducted to study the effect of orlistat on jejunal cholesterol absorption. Inhibition kinetic parameters were calculated from in vitro inhibition studies using Caco2 and NPC1L1 transfected cell lines. The in situ studies demonstrated that intestinal perfusion of orlistat (100µM) was able to reduce cholesterol absorption by three-fold when compared to control (i.e. in the absence of orlistat, Pabsorption of cholesterol, we demonstrated for the first time that orlistat limits cholesterol absorption by the inhibition of NPC1L1 transport protein. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. A Plasmodium falciparum copper-binding membrane protein with copper transport motifs

    Directory of Open Access Journals (Sweden)

    Choveaux David L

    2012-11-01

    Full Text Available Abstract Background Copper is an essential catalytic co-factor for metabolically important cellular enzymes, such as cytochrome-c oxidase. Eukaryotic cells acquire copper through a copper transport protein and distribute intracellular copper using molecular chaperones. The copper chelator, neocuproine, inhibits Plasmodium falciparum ring-to-trophozoite transition in vitro, indicating a copper requirement for malaria parasite development. How the malaria parasite acquires or secretes copper still remains to be fully elucidated. Methods PlasmoDB was searched for sequences corresponding to candidate P. falciparum copper-requiring proteins. The amino terminal domain of a putative P. falciparum copper transport protein was cloned and expressed as a maltose binding fusion protein. The copper binding ability of this protein was examined. Copper transport protein-specific anti-peptide antibodies were generated in chickens and used to establish native protein localization in P. falciparum parasites by immunofluorescence microscopy. Results Six P. falciparum copper-requiring protein orthologs and a candidate P. falciparum copper transport protein (PF14_0369, containing characteristic copper transport protein features, were identified in PlasmoDB. The recombinant amino terminal domain of the transport protein bound reduced copper in vitro and within Escherichia coli cells during recombinant expression. Immunolocalization studies tracked the copper binding protein translocating from the erythrocyte plasma membrane in early ring stage to a parasite membrane as the parasites developed to schizonts. The protein appears to be a PEXEL-negative membrane protein. Conclusion Plasmodium falciparum parasites express a native protein with copper transporter characteristics that binds copper in vitro. Localization of the protein to the erythrocyte and parasite plasma membranes could provide a mechanism for the delivery of novel anti-malarial compounds.

  14. Prediction of membrane transport proteins and their substrate specificities using primary sequence information.

    Directory of Open Access Journals (Sweden)

    Nitish K Mishra

    Full Text Available Membrane transport proteins (transporters move hydrophilic substrates across hydrophobic membranes and play vital roles in most cellular functions. Transporters represent a diverse group of proteins that differ in topology, energy coupling mechanism, and substrate specificity as well as sequence similarity. Among the functional annotations of transporters, information about their transporting substrates is especially important. The experimental identification and characterization of transporters is currently costly and time-consuming. The development of robust bioinformatics-based methods for the prediction of membrane transport proteins and their substrate specificities is therefore an important and urgent task.Support vector machine (SVM-based computational models, which comprehensively utilize integrative protein sequence features such as amino acid composition, dipeptide composition, physico-chemical composition, biochemical composition, and position-specific scoring matrices (PSSM, were developed to predict the substrate specificity of seven transporter classes: amino acid, anion, cation, electron, protein/mRNA, sugar, and other transporters. An additional model to differentiate transporters from non-transporters was also developed. Among the developed models, the biochemical composition and PSSM hybrid model outperformed other models and achieved an overall average prediction accuracy of 76.69% with a Mathews correlation coefficient (MCC of 0.49 and a receiver operating characteristic area under the curve (AUC of 0.833 on our main dataset. This model also achieved an overall average prediction accuracy of 78.88% and MCC of 0.41 on an independent dataset.Our analyses suggest that evolutionary information (i.e., the PSSM and the AAIndex are key features for the substrate specificity prediction of transport proteins. In comparison, similarity-based methods such as BLAST, PSI-BLAST, and hidden Markov models do not provide accurate predictions

  15. Prediction of membrane transport proteins and their substrate specificities using primary sequence information.

    Science.gov (United States)

    Mishra, Nitish K; Chang, Junil; Zhao, Patrick X

    2014-01-01

    Membrane transport proteins (transporters) move hydrophilic substrates across hydrophobic membranes and play vital roles in most cellular functions. Transporters represent a diverse group of proteins that differ in topology, energy coupling mechanism, and substrate specificity as well as sequence similarity. Among the functional annotations of transporters, information about their transporting substrates is especially important. The experimental identification and characterization of transporters is currently costly and time-consuming. The development of robust bioinformatics-based methods for the prediction of membrane transport proteins and their substrate specificities is therefore an important and urgent task. Support vector machine (SVM)-based computational models, which comprehensively utilize integrative protein sequence features such as amino acid composition, dipeptide composition, physico-chemical composition, biochemical composition, and position-specific scoring matrices (PSSM), were developed to predict the substrate specificity of seven transporter classes: amino acid, anion, cation, electron, protein/mRNA, sugar, and other transporters. An additional model to differentiate transporters from non-transporters was also developed. Among the developed models, the biochemical composition and PSSM hybrid model outperformed other models and achieved an overall average prediction accuracy of 76.69% with a Mathews correlation coefficient (MCC) of 0.49 and a receiver operating characteristic area under the curve (AUC) of 0.833 on our main dataset. This model also achieved an overall average prediction accuracy of 78.88% and MCC of 0.41 on an independent dataset. Our analyses suggest that evolutionary information (i.e., the PSSM) and the AAIndex are key features for the substrate specificity prediction of transport proteins. In comparison, similarity-based methods such as BLAST, PSI-BLAST, and hidden Markov models do not provide accurate predictions for the

  16. Regulatory pathways for ATP-binding cassette transport proteins in kidney proximal tubules.

    Science.gov (United States)

    Masereeuw, Rosalinde; Russel, Frans G M

    2012-12-01

    The ATP-binding cassette transport proteins (ABC transporters) represent important determinants of drug excretion. Protective or excretory tissues where these transporters mediate substrate efflux include the kidney proximal tubule. Regulation of the transport proteins in this tissue requires elaborate signaling pathways, including genetic, epigenetic, nuclear receptor mediated, posttranscriptional gene regulation involving microRNAs, and non-genomic (kinases) pathways triggered by hormones and/or growth factors. This review discusses current knowledge on regulatory pathways for ABC transporters in kidney proximal tubules, with a main focus on P-glycoprotein, multidrug resistance proteins 2 and 4, and breast cancer resistance protein. Insight in these processes is of importance because variations in transporter activity due to certain (disease) conditions could lead to significant changes in drug efficacy or toxicity.

  17. Analysis of Nanobody-Epitope Interactions in Living Cells via Quantitative Protein Transport Assays.

    Science.gov (United States)

    Früholz, Simone; Pimpl, Peter

    2017-01-01

    Over the past few decades, quantitative protein transport analyses have been used to elucidate the sorting and transport of proteins in the endomembrane system of plants. Here, we have applied our knowledge about transport routes and the corresponding sorting signals to establish an in vivo system for testing specific interactions between soluble proteins.Here, we describe the use of quantitative protein transport assays in tobacco mesophyll protoplasts to test for interactions occurring between a GFP-binding nanobody and its GFP epitope. For this, we use a secreted GFP-tagged α-amylase as a reporter together with a vacuolar-targeted RFP-tagged nanobody. The interaction between these proteins is then revealed by a transport alteration of the secretory reporter due to the interaction-triggered attachment of the vacuolar sorting signal.

  18. MLN64 induces mitochondrial dysfunction associated with increased mitochondrial cholesterol content

    Directory of Open Access Journals (Sweden)

    Elisa Balboa

    2017-08-01

    Full Text Available MLN64 is a late endosomal cholesterol-binding membrane protein that has been implicated in cholesterol transport from endosomal membranes to the plasma membrane and/or mitochondria, in toxin-induced resistance, and in mitochondrial dysfunction. Down-regulation of MLN64 in Niemann-Pick C1 deficient cells decreased mitochondrial cholesterol content, suggesting that MLN64 functions independently of NPC1. However, the role of MLN64 in the maintenance of endosomal cholesterol flow and intracellular cholesterol homeostasis remains unclear. We have previously described that hepatic MLN64 overexpression increases liver cholesterol content and induces liver damage. Here, we studied the function of MLN64 in normal and NPC1-deficient cells and we evaluated whether MLN64 overexpressing cells exhibit alterations in mitochondrial function. We used recombinant-adenovirus-mediated MLN64 gene transfer to overexpress MLN64 in mouse liver and hepatic cells; and RNA interference to down-regulate MLN64 in NPC1-deficient cells. In MLN64-overexpressing cells, we found increased mitochondrial cholesterol content and decreased glutathione (GSH levels and ATPase activity. Furthermore, we found decreased mitochondrial membrane potential and mitochondrial fragmentation and increased mitochondrial superoxide levels in MLN64-overexpressing cells and in NPC1-deficient cells. Consequently, MLN64 expression was increased in NPC1-deficient cells and reduction of its expression restore mitochondrial membrane potential and mitochondrial superoxide levels. Our findings suggest that MLN64 overexpression induces an increase in mitochondrial cholesterol content and consequently a decrease in mitochondrial GSH content leading to mitochondrial dysfunction. In addition, we demonstrate that MLN64 expression is increased in NPC cells and plays a key role in cholesterol transport into the mitochondria.

  19. MLN64 induces mitochondrial dysfunction associated with increased mitochondrial cholesterol content.

    Science.gov (United States)

    Balboa, Elisa; Castro, Juan; Pinochet, María-José; Cancino, Gonzalo I; Matías, Nuria; Sáez, P J; Martínez, Alexis; Álvarez, Alejandra R; Garcia-Ruiz, Carmen; Fernandez-Checa, José C; Zanlungo, Silvana

    2017-08-01

    MLN64 is a late endosomal cholesterol-binding membrane protein that has been implicated in cholesterol transport from endosomal membranes to the plasma membrane and/or mitochondria, in toxin-induced resistance, and in mitochondrial dysfunction. Down-regulation of MLN64 in Niemann-Pick C1 deficient cells decreased mitochondrial cholesterol content, suggesting that MLN64 functions independently of NPC1. However, the role of MLN64 in the maintenance of endosomal cholesterol flow and intracellular cholesterol homeostasis remains unclear. We have previously described that hepatic MLN64 overexpression increases liver cholesterol content and induces liver damage. Here, we studied the function of MLN64 in normal and NPC1-deficient cells and we evaluated whether MLN64 overexpressing cells exhibit alterations in mitochondrial function. We used recombinant-adenovirus-mediated MLN64 gene transfer to overexpress MLN64 in mouse liver and hepatic cells; and RNA interference to down-regulate MLN64 in NPC1-deficient cells. In MLN64-overexpressing cells, we found increased mitochondrial cholesterol content and decreased glutathione (GSH) levels and ATPase activity. Furthermore, we found decreased mitochondrial membrane potential and mitochondrial fragmentation and increased mitochondrial superoxide levels in MLN64-overexpressing cells and in NPC1-deficient cells. Consequently, MLN64 expression was increased in NPC1-deficient cells and reduction of its expression restore mitochondrial membrane potential and mitochondrial superoxide levels. Our findings suggest that MLN64 overexpression induces an increase in mitochondrial cholesterol content and consequently a decrease in mitochondrial GSH content leading to mitochondrial dysfunction. In addition, we demonstrate that MLN64 expression is increased in NPC cells and plays a key role in cholesterol transport into the mitochondria. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  20. The cholesterol space of the rat

    International Nuclear Information System (INIS)

    Chevallier, F.

    1959-01-01

    The experiments consisted in feeding daily to rats the same mass of radioactive cholesterol, over variable time intervals. From the evolution of the specific radioactivity of cholesterol carbon-14 in the organs as a function of time, information relative to the transport of cholesterol in the organism may be obtained. 1) The cholesterol space, defined as the group of molecules capable of being transferred from the organs into the serum and vice versa, represents at the most 50 per cent of the total cholesterol of the adult rat. 2) The incessant interchange between the tissual and the serum cholesterol renews entirely or for the most part the cholesterol molecules contained in the following organs: spleen, heart, adipose tissue, suprarenal glands, lungs, bone marrow, liver, erythrocytes. For a second group of organs: skin, testicles, kidneys, colon, bones, muscles, only a fraction of their cholesterol is renewable by this process. No transfer can be detected at the level of the brain. 3) The relative speeds of the various means of appearance (absorption, synthesis) and disappearance (excretion, transformation) of the cholesterol from its space are such that a stationary isotopic state is established around the eighth day, when the animal absorbs 5 milligrams of radioactive cholesterol daily. (author) [fr

  1. The UPS and downs of cholesterol homeostasis

    NARCIS (Netherlands)

    Sharpe, Laura J.; Cook, Emma C. L.; Zelcer, Noam; Brown, Andrew J.

    2014-01-01

    An emerging theme in the regulation of cholesterol homeostasis is the role of the ubiquitin proteasome system (UPS), through which proteins are ubiquitylated and then degraded in response to specific signals. The UPS controls all aspects of cholesterol metabolism including its synthesis, uptake, and

  2. Mechanism of transfer of LDL-derived free cholesterol to HDL subfractions in human plasma

    International Nuclear Information System (INIS)

    Miida, T.; Fielding, C.J.; Fielding, P.E.

    1990-01-01

    The transfer of [ 3 H]cholesterol in low-density lipoprotein (LDL) to different high-density lipoprotein (HDL) species in native human plasma was determined by using nondenaturing two-dimensional electrophoresis. Transfer from LDL had a t 1/2 at 37 degree C of 51 ± 8 min and an activation energy of 18.0 kCal mol -1 . There was unexpected specificity among HDL species as acceptors of LDL-derived labeled cholesterol. The largest fraction of the major α-migrating class (HDL 2b ) was the major initial acceptor of LDL-derived cholesterol. Kinetic analysis indicated a rapid secondary transfer from HDL 2b to smaller αHDL (particularly HDL 3 ) driven enzymatically by the lecithin-cholesterol acyltransferase reaction. Rates of transfer among αHDL were most rapid from the largest αHDL fraction (HDL 2b ), suggesting possible protein-mediated facilitation. Simultaneous measurements of the transport of LDL-derived and cell-derived isotopic cholesterol indicated that the former preferably utilized the αHDL pathyway, with little label in pre-βHDL. The same experiments confirmed earlier data that cell-derived cholesterol is preferentially channeled through pre-βHDL. The authors suggest that the functional heterogeneity of HDL demonstrated here includes the ability to independently process cell- and LDL-derived free cholesterol

  3. LDL-Cholesterol Increases the Transcytosis of Molecules through Endothelial Monolayers.

    Science.gov (United States)

    Magalhaes, Ana; Matias, Inês; Palmela, Inês; Brito, Maria Alexandra; Dias, Sérgio

    2016-01-01

    Cholesterol has been identified as a causative factor in numerous pathologies including atherosclerosis and cancer. One of the frequent effects of elevated cholesterol levels in humans is the compromise of endothelial function due to activation of pro-inflammatory signalling pathways. While the mechanisms involved in endothelial activation by cholesterol during an inflammatory response are well established, less is known about the mechanisms by which cholesterol may affect endothelial barrier function, which were the subject of the present study. Here we show that low density lipoprotein (LDL) increases the permeability of endothelial monolayers to high molecular weight dextrans in an LDL receptor and cholesterol-dependent manner. The increased permeability seen upon LDL treatment was not caused by disruption of cell-to-cell junctions as determined by a normal localization of VE-Cadherin and ZO-1 proteins, and no major alterations in transendothelial electrical resistance or permeability to fluorescein. We show instead that LDL increases the level of high molecular weight transcytosis and that this occurs in an LDL receptor, cholesterol and caveolae-dependent way. Our findings contribute to our understanding of the systemic pathological effects of elevated cholesterol and the transport of cargo through endothelial monolayers.

  4. SDS-assisted protein transport through solid-state nanopores.

    Science.gov (United States)

    Restrepo-Pérez, Laura; John, Shalini; Aksimentiev, Aleksei; Joo, Chirlmin; Dekker, Cees

    2017-08-17

    Using nanopores for single-molecule sequencing of proteins - similar to nanopore-based sequencing of DNA - faces multiple challenges, including unfolding of the complex tertiary structure of the proteins and enforcing their unidirectional translocation through nanopores. Here, we combine molecular dynamics (MD) simulations with single-molecule experiments to investigate the utility of SDS (Sodium Dodecyl Sulfate) to unfold proteins for solid-state nanopore translocation, while simultaneously endowing them with a stronger electrical charge. Our simulations and experiments prove that SDS-treated proteins show a considerable loss of the protein structure during the nanopore translocation. Moreover, SDS-treated proteins translocate through the nanopore in the direction prescribed by the electrophoretic force due to the negative charge impaired by SDS. In summary, our results suggest that SDS causes protein unfolding while facilitating protein translocation in the direction of the electrophoretic force; both characteristics being advantageous for future protein sequencing applications using solid-state nanopores.

  5. Assessment of modes of action and efficacy of plasma cholesterol-lowering drugs : measurement of cholesterol absorption, cholesterol synthesis and bile acid synthesis and turnover using novel stable isotope techniques

    NARCIS (Netherlands)

    Stellaard, Frans; Kuipers, Folkert

    Several processes are involved in control of plasma cholesterol levels, e.g., intestinal cholesterol absorption, endogenous cholesterol synthesis and transport and bile acid synthesis. Adaptation of either of these processes allows the body to adapt to changes in dietary cholesterol intake.

  6. Intracellular transport of recombinant coronavirus spike proteins: implications for virus assembly

    NARCIS (Netherlands)

    Horzinek, M.C.; Vennema, H.; Heijnen, L.; Zijderveld, A.; Spaan, W.J.M.

    1990-01-01

    Coronavirus spike protein genes were expressed in vitro by using the recombinant vaccinia virus expression system. Recombinant spike proteins were expressed at the cell surface and induced cell fusion in a host-cell-dependent fashion. The intracellular transport of recombinant spike proteins was

  7. Binding proteins enhance specific uptake rate by increasing the substrate-transporter encounter rate.

    NARCIS (Netherlands)

    Bosdriesz, E.; Magnúsdóttir, S.; Bruggeman, F.J.; Teusink, B.; Molenaar, D.

    2015-01-01

    Microorganisms rely on binding-protein assisted, active transport systems to scavenge for scarce nutrients. Several advantages of using binding proteins in such uptake systems have been proposed. However, a systematic, rigorous and quantitative analysis of the function of binding proteins is

  8. Alcohol consumption, TaqIB polymorphism of cholesteryl ester transfer protein, high-density lipoprotein cholesterol, and risk of coronary heart disease in men and women.

    Science.gov (United States)

    Jensen, Majken K; Mukamal, Kenneth J; Overvad, Kim; Rimm, Eric B

    2008-01-01

    To investigate whether a common polymorphism in the cholesteryl ester transfer protein (CETP) gene modifies the relationship of alcohol intake with high-density lipoprotein cholesterol (HDL-C) and risk of coronary heart disease (CHD). Parallel nested case-control studies among women [Nurses' Health Study (NHS)] and men [Health Professionals Follow-up Study (HPFS)] where 246 women and 259 men who developed incident CHD were matched to controls (1:2) on age and smoking. The TaqIB variant and alcohol consumption were associated with higher HDL-C, with the most pronounced effects of alcohol among B2 carriers. In the NHS we did not find an inverse association between alcohol and CHD in B2 non-carriers (P trend: 0.5), but did among B2 carriers (P trend alcohol with HDL-C levels was modified by CETP TaqIB2 carrier status, and there was also a suggestion of a gene-environment interaction on the risk of CHD.

  9. Fast axonal transport of labeled proteins in motoneurons of exercise-trained rats

    International Nuclear Information System (INIS)

    Jasmin, B.J.; Lavoie, P.A.; Gardiner, P.F.

    1988-01-01

    In this study, the fast orthograde axonal transport of radiolabeled proteins was measured to determine the effects of endurance-running training on transport velocity and amounts of transported proteins in rat sciatic motoneurons. Female rats were subjected to a progressive running-training program for 10-12 wk. Twenty-four hours after the last training session, rats underwent right L4-L5 dorsal root ganglionectomy. The next day, 20 microCi of [3H]leucine was injected bilaterally in the vicinity of the motoneuronal cell bodies supplying the sciatic nerve, to study axonal transport parameters. Results showed that peak and average transport velocities of labeled proteins were significantly (P less than 0.05) increased by 22 and 29%, respectively, in the deafferented nerves of the runners as compared with controls. Moreover, the amount of total transported protein-bound radioactivity was increased in both left (40%) and right (37%) sciatic nerves of the runners. An exhaustive exercise session reduced (P less than 0.05) peak displacement (8%) and total transported protein-bound radioactivity (36%) in the sciatic nerves of control rats, whereas no changes were noticed in trained animals. The data suggest that chronic endurance running induces significant adaptations in the fast axonal transport of labeled proteins

  10. Efficient retrograde transport of pseudorabies virus within neurons requires local protein synthesis in axons.

    Science.gov (United States)

    Koyuncu, Orkide O; Perlman, David H; Enquist, Lynn W

    2013-01-16

    After replicating in epithelial cells, alphaherpesviruses such as pseudorabies virus (PRV) invade axons of peripheral nervous system neurons and undergo retrograde transport toward the distant cell bodies. Although several viral proteins engage molecular motors to facilitate transport, the initial steps and neuronal responses to infection are poorly understood. Using compartmented neuron cultures to physically separate axon infection from cell bodies, we found that PRV infection induces local protein synthesis in axons, including proteins involved in cytoskeletal remodeling, intracellular trafficking, signaling, and metabolism. This rapid translation of axonal mRNAs is required for efficient PRV retrograde transport and infection of cell bodies. Furthermore, induction of axonal damage, which also induces local protein synthesis, prior to infection reduces virion trafficking, suggesting that host damage signals and virus particles compete for retrograde transport. Thus, similar to axonal damage, virus infection induces local protein translation in axons, and viruses likely exploit this response for invasion. Copyright © 2013 Elsevier Inc. All rights reserved.

  11. [Effect of ferulic acid on cholesterol efflux in macrophage foam cell formation and potential mechanism].

    Science.gov (United States)

    Chen, Fu-xin; Wang, Lian-kai

    2015-02-01

    The formation of macrophage-derived foam cells is a typical feature of atherosclerosis (AS). Reverse cholesterol efflux (RCT) is one of important factors for the formation of macrophage foam cells. In this study, macrophage form cells were induced by oxidized low density lipoprotein (ox-LDL) and then treated with different concentrations of ferulic acid, so as to observe the effect of ferulic acid on the intracellular lipid metabolism in the ox-LDL-induced macrophage foam cell formation, the cholesterol efflux and the mRNA expression and protein levels of ATP binding cassette transporter A1 (ABCA1) and ATP binding cassette transporter G1 (ABCG1) that mediate cholesterol efflux, and discuss the potential mechanism of ferulic acid in resisting AS. According to the findings, compared with the control group, the ox-LDL-treated group showed significant increase in intracellular lipid content, especially for the cholesterol content; whereas the intracellular lipid accumulation markedly decreased, after the treatment with ferulic acid. The data also demonstrated that the mRNA and protein expressions of ABCA1 and ABCG1 significantly increased after macrophage foam cells were treated with different concentrations of ferulic acid. In summary, ferulic acid may show the anti-atherosclerosis effect by increasing the surface ABCA1 and ABCG1 expressions of macrophage form cells and promoting cholesterol efflux.

  12. The Arabidopsis NPF3 protein is a GA transporter

    DEFF Research Database (Denmark)

    Tal, Iris; Zhang, Yi; Jørgensen, Morten Egevang

    2016-01-01

    Gibberellins (GAs) are plant hormones that promote a wide range of developmental processes. While GA signalling is well understood, little is known about how GA is transported or how GA distribution is regulated. Here we utilize fluorescently labelled GAs (GA-Fl) to screen for Arabidopsis mutants...... deficient in GA transport. We show that the NPF3 transporter efficiently transports GA across cell membranes in vitro and GA-Fl in vivo. NPF3 is expressed in root endodermis and repressed by GA. NPF3 is targeted to the plasma membrane and subject to rapid BFA-dependent recycling. We show that abscisic acid...... (ABA), an antagonist of GA, is also transported by NPF3 in vitro. ABA promotes NPF3 expression and GA-Fl uptake in plants. On the basis of these results, we propose that GA distribution and activity in Arabidopsis is partly regulated by NPF3 acting as an influx carrier and that GA-ABA interaction may...

  13. Isolation of Arabidopsis thylakoid membranes and their use for in vitro protein insertion or transport assays.

    Science.gov (United States)

    Bals, Thomas; Schünemann, Danja

    2011-01-01

    This chapter focuses on the techniques of chloroplast isolation; their fractionation into envelopes, stroma, and thylakoids; and their further use for in vitro protein transport assays. In addition to the isolation of thylakoids, this chapter also describes the experimental steps of both protein translocation across the thylakoid membrane and protein integration into the membrane. Protein translocation and integration can be analysed by the radioactive labelling of substrate proteins using an in vitro transcription and translation system. The translocated or integrated proteins can then be detected by autoradiography. Our protocol allows the analysis of these transport systems in wild-type Arabidopsis or mutants that lack or overexpress soluble or membrane transport factors that could be of potential interest.

  14. Omega 3 fatty acids chemosensitize multidrug resistant colon cancer cells by down-regulating cholesterol synthesis and altering detergent resistant membranes composition.

    OpenAIRE

    Gelsomino, G; Gazzano, E

    2013-01-01

    Background The activity of P-glycoprotein (Pgp) and multidrug resistance related protein 1 (MRP1), two membrane transporters involved in multidrug resistance of colon cancer, is increased by high amounts of cholesterol in plasma membrane and detergent resistant membranes (DRMs). It has never been investigated whether omega 3 polyunsatured fatty acids (PUFAs), which modulate cholesterol homeostasis in dyslipidemic syndromes and have chemopreventive effects in colon cancer, may affect the respo...

  15. Active zone proteins are transported via distinct mechanisms regulated by Par-1 kinase.

    Directory of Open Access Journals (Sweden)

    Kara R Barber

    2017-02-01

    Full Text Available Disruption of synapses underlies a plethora of neurodevelopmental and neurodegenerative disease. Presynaptic specialization called the active zone plays a critical role in the communication with postsynaptic neuron. While the role of many proteins at the active zones in synaptic communication is relatively well studied, very little is known about how these proteins are transported to the synapses. For example, are there distinct mechanisms for the transport of active zone components or are they all transported in the same transport vesicle? Is active zone protein transport regulated? In this report we show that overexpression of Par-1/MARK kinase, a protein whose misregulation has been implicated in Autism spectrum disorders (ASDs and neurodegenerative disorders, lead to a specific block in the transport of an active zone protein component- Bruchpilot at Drosophila neuromuscular junctions. Consistent with a block in axonal transport, we find a decrease in number of active zones and reduced neurotransmission in flies overexpressing Par-1 kinase. Interestingly, we find that Par-1 acts independently of Tau-one of the most well studied substrates of Par-1, revealing a presynaptic function for Par-1 that is independent of Tau. Thus, our study strongly suggests that there are distinct mechanisms that transport components of active zones and that they are tightly regulated.

  16. A Short Synthetic Peptide Mimetic of Apolipoprotein A1 Mediates Cholesterol and Globotriaosylceramide Efflux from Fabry Fibroblasts.

    Science.gov (United States)

    Schueler, Ulrike; Kaneski, Christine; Remaley, Alan; Demosky, Stephen; Dwyer, Nancy; Blanchette-Mackie, Joan; Hanover, John; Brady, Roscoe

    2016-01-01

    Fabry disease is an X-linked sphingolipid storage disorder caused by a deficiency of the lysosomal enzyme α-galactosidase A (AGA, EC 3.2.1.22) resulting in the intracellular accumulation of globotriaosylceramide (Gb3). We found that Gb3 storage also correlates with accumulation of endosomal-lysosomal cholesterol in Fabry fibroblasts. This cholesterol accumulation may contribute to the phenotypic pathology of Fabry disease by slowing endosomal-lysosomal trafficking. We found that LDL receptor expression is not downregulated in Fabry fibroblasts resulting in accumulation of both cholesterol and Gb3. 5A-Palmitoyl oleoyl-phosphatidylcholine (5AP) is a phospholipid complex containing a short synthetic peptide that mimics apolipoprotein A1, the main protein component of high-density lipoprotein (HDL) that mediates the efflux of cholesterol from cells via the ATP-binding cassette transporter. We used 5AP and HDL to remove cholesterol from Fabry fibroblasts to examine the fate of accumulated cellular Gb3. Using immunostaining techniques, we found that 5AP is highly effective for depleting cholesterol and Gb3 in these cells. 5AP restores the ApoA-1-mediated cholesterol efflux leading to mobilization of cholesterol and reduction of Gb3 in Fabry fibroblasts.

  17. High Density Lipoprotein Cholesterol Increasing Therapy: The Unmet Cardiovascular Need

    Science.gov (United States)

    Cimmino, Giovanni; Ciccarelli, Giovanni; Morello, Alberto; Ciccarelli, Michele; Golino, Paolo

    2015-01-01

    Despite aggressive strategies are now available to reduce LDL-cholesterol, the risk of cardiovascular events in patients with coronary artery disease remains substantial. Several preclinical and clinical studies have shown that drug therapy ultimately leads to a regression of the angiographic lesions but also results in a reduction in cardiovascular events. The dramatic failure of clinical trials evaluating the cholesterol ester transfer protein (CEPT) inhibitors, torcetrapib and dalcetrapib, has led to considerable doubt about the value of the current strategy to raise high-density lipoprotein cholesterol (HDL-C) as a treatment for cardiovascular disease. These clinical results, as well as animal studies, have revealed the complexity of HDL metabolism, assessing a more important role of functional quality compared to circulating quantity of HDL. As a result, HDL-based therapeutic interventions that maintain or enhance HDL functionality, such as improving its main property, the reverse cholesterol transport, require closer investigation. In this review, we will discuss HDL metabolism and function, clinical-trial data available for HDL-raising agents, and potential strategies for future HDL-based therapies. PMID:26535185

  18. A cell-based method for screening RNA-protein interactions: identification of constitutive transport element-interacting proteins.

    Directory of Open Access Journals (Sweden)

    Robert L Nakamura

    Full Text Available We have developed a mammalian cell-based screening platform to identify proteins that assemble into RNA-protein complexes. Based on Tat-mediated activation of the HIV LTR, proteins that interact with an RNA target elicit expression of a GFP reporter and are captured by fluorescence activated cell sorting. This "Tat-hybrid" screening platform was used to identify proteins that interact with the Mason Pfizer monkey virus (MPMV constitutive transport element (CTE, a structured RNA hairpin that mediates the transport of unspliced viral mRNAs from the nucleus to the cytoplasm. Several hnRNP-like proteins, including hnRNP A1, were identified and shown to interact with the CTE with selectivity in the reporter system comparable to Tap, a known CTE-binding protein. In vitro gel shift and pull-down assays showed that hnRNP A1 is able to form a complex with the CTE and Tap and that the RGG domain of hnRNP A1 mediates binding to Tap. These results suggest that hnRNP-like proteins may be part of larger export-competent RNA-protein complexes and that the RGG domains of these proteins play an important role in directing these binding events. The results also demonstrate the utility of the screening platform for identifying and characterizing new components of RNA-protein complexes.

  19. Transport of soluble proteins through the Golgi occurs by diffusion via continuities across cisternae

    Science.gov (United States)

    Beznoussenko, Galina V; Parashuraman, Seetharaman; Rizzo, Riccardo; Polishchuk, Roman; Martella, Oliviano; Di Giandomenico, Daniele; Fusella, Aurora; Spaar, Alexander; Sallese, Michele; Capestrano, Maria Grazia; Pavelka, Margit; Vos, Matthijn R; Rikers, Yuri GM; Helms, Volkhard; Mironov, Alexandre A; Luini, Alberto

    2014-01-01

    The mechanism of transport through the Golgi complex is not completely understood, insofar as no single transport mechanism appears to account for all of the observations. Here, we compare the transport of soluble secretory proteins (albumin and α1-antitrypsin) with that of supramolecular cargoes (e.g., procollagen) that are proposed to traverse the Golgi by compartment progression–maturation. We show that these soluble proteins traverse the Golgi much faster than procollagen while moving through the same stack. Moreover, we present kinetic and morphological observations that indicate that albumin transport occurs by diffusion via intercisternal continuities. These data provide evidence for a transport mechanism that applies to a major class of secretory proteins and indicate the co-existence of multiple intra-Golgi trafficking modes. DOI: http://dx.doi.org/10.7554/eLife.02009.001 PMID:24867214

  20. Dietary cholesterol - the role of eggs in the prudent diet

    African Journals Online (AJOL)

    cholesterol levels and coronary heart disease is often used to justify a ... Department of Nutrition, Potchefstroom University, N.-W. H. H. Vorster, D.SC. C. S. Venter, D.SC. Dietary cholesterol - the role of eggs in the prudent diet. Department of Human .... cholesterol/MJ;. 240 mg/day) without changes in protein or fat intake, the.

  1. Controlling Cholesterol with Statins

    Science.gov (United States)

    ... For Consumers Home For Consumers Consumer Updates Controlling Cholesterol with Statins Share Tweet Linkedin Pin it More ... not, the following tips can help keep your cholesterol in check: Talk with your healthcare provider about ...

  2. Cholesterol - drug treatment

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/patientinstructions/000314.htm Cholesterol - drug treatment To use the sharing features on ... treatment; Hardening of the arteries - statin Statins for Cholesterol Statins reduce your risk of heart disease, stroke, ...

  3. LDL: The "Bad" Cholesterol

    Science.gov (United States)

    ... waxy, fat-like substance that's found in all the cells in your body. Your liver makes cholesterol, ... stands for low-density lipoproteins. It is called the "bad" cholesterol because a high LDL level leads ...

  4. Polyunsaturated fatty acyl-coenzyme As are inhibitors of cholesterol biosynthesis in zebrafish and mice

    Directory of Open Access Journals (Sweden)

    Santhosh Karanth

    2013-11-01

    Lipid disorders pose therapeutic challenges. Previously we discovered that mutation of the hepatocyte β-hydroxybutyrate transporter Slc16a6a in zebrafish causes hepatic steatosis during fasting, marked by increased hepatic triacylglycerol, but not cholesterol. This selective diversion of trapped ketogenic carbon atoms is surprising because acetate and acetoacetate can exit mitochondria and can be incorporated into both fatty acids and cholesterol in normal hepatocytes. To elucidate the mechanism of this selective diversion of carbon atoms to fatty acids, we fed wild-type and slc16a6a mutant animals high-protein ketogenic diets. We find that slc16a6a mutants have decreased activity of the rate-limiting enzyme of cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (Hmgcr, despite increased Hmgcr protein abundance and relative incorporation of mevalonate into cholesterol. These observations suggest the presence of an endogenous Hmgcr inhibitor. We took a candidate approach to identify such inhibitors. First, we found that mutant livers accumulate multiple polyunsaturated fatty acids (PUFAs and PUFA-CoAs, and we showed that human HMGCR is inhibited by PUFA-CoAs in vitro. Second, we injected mice with an ethyl ester of the PUFA eicosapentaenoic acid and observed an acute decrease in hepatic Hmgcr activity, without alteration in Hmgcr protein abundance. These results elucidate a mechanism for PUFA-mediated cholesterol lowering through direct inhibition of Hmgcr.

  5. SDS-assisted protein transport through solid-state nanopores

    NARCIS (Netherlands)

    Restrepo Perez, L.; John, Shalini; Aksimentiev, Aleksei; Joo, C.; Dekker, C.

    2017-01-01

    Using nanopores for single-molecule sequencing of proteins – similar to nanopore-based sequencing of DNA – faces multiple challenges, including unfolding of the complex tertiary structure of the proteins and enforcing their unidirectional translocation through nanopores. Here, we combine molecular

  6. Nuclear transport factor directs localization of protein synthesis during mitosis

    NARCIS (Netherlands)

    Bogaart, Geert van den; Meinema, Anne C.; Krasnikov, Viktor; Veenhoff, Liesbeth M.; Poolman, Bert

    Export of messenger RNA from the transcription site in the nucleus and mRNA targeting to the translation site in the cytoplasm are key regulatory processes in protein synthesis. In yeast, the mRNA-binding proteins Nab2p and Nab4p/Hrp1p accompany transcripts to their translation site, where the

  7. Protein transport across the small intestine in food allergy

    NARCIS (Netherlands)

    Reitsma, M.; Westerhout, J.; Wichers, H.J.; Wortelboer, H.M.; Verhoeckx, K.C.M.

    2014-01-01

    In view of the imminent deficiency of protein sources for human consumption in the near future, new protein sources need to be identified. However, safety issues such as the risk of allergenicity are often a bottleneck, due to the absence of predictive, validated and accepted methods for risk

  8. Protein transport across the small intestine in food hypersensitivity

    NARCIS (Netherlands)

    Reitsma, M.; Westerhout, J.; Wichers, H.J.; Wortelboer, H.; Verhoeckx, K.C.M.

    2014-01-01

    In view of the imminent deficiency of protein sources for human consumption in the near future, new protein sources need to be identified. However, safety issues such as the risk of allergenicity are often a bottleneck, due to the absence of predictive, validated and accepted methods for risk

  9. Communication Maps: Exploring Energy Transport through Proteins and Water

    Czech Academy of Sciences Publication Activity Database

    Agbo, J. K.; Gnanasekaran, Ramachandran; Leitner, D. M.

    2014-01-01

    Roč. 54, 8/9 (2014), s. 1065-1073 ISSN 0021-2148 Institutional support: RVO:61388963 Keywords : energy transfer * heme proteins * hydrogen bonds * molecular modeling * protein models Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.221, year: 2014

  10. High-density lipoprotein cholesterol: How High

    Directory of Open Access Journals (Sweden)

    G Rajagopal

    2012-01-01

    Full Text Available The high-density lipoprotein cholesterol (HDL-C is considered anti-atherogenic good cholesterol. It is involved in reverse transport of lipids. Epidemiological studies have found inverse relationship of HDL-C and coronary heart disease (CHD risk. When grouped according to HDL-C, subjects having HDL-C more than 60 mg/dL had lesser risk of CHD than those having HDL-C of 40-60 mg/dL, who in turn had lesser risk than those who had HDL-C less than 40 mg/dL. No upper limit for beneficial effect of HDL-C on CHD risk has been identified. The goals of treating patients with low HDL-C have not been firmly established. Though many drugs are known to improve HDL-C concentration, statins are proven to improve CHD risk and mortality. Cholesteryl ester transfer protein (CETP is involved in metabolism of HDL-C and its inhibitors are actively being screened for clinical utility. However, final answer is still awaited on CETP-inhibitors.

  11. Mitochondrial damage and cholesterol storage in human hepatocellular carcinoma cells with silencing of UBIAD1 gene expression

    Directory of Open Access Journals (Sweden)

    Carlos R. Morales

    2014-01-01

    Full Text Available Heterozygous mutations in the UBIAD1 gene cause Schnyder corneal dystrophy characterized by abnormal cholesterol and phospholipid deposits in the cornea. Ubiad1 protein was recently identified as Golgi prenyltransferase responsible for biosynthesis of vitamin K2 and CoQ10, a key protein in the mitochondrial electron transport chain. Our study shows that silencing UBIAD1 in cultured human hepatocellular carcinoma cells causes dramatic morphological changes and cholesterol storage in the mitochondria, emphasizing an important role of UBIAD1 in mitochondrial function.

  12. Potassium-transporting proteins in skeletal muscle: cellular location and fiber-type differences

    DEFF Research Database (Denmark)

    Kristensen, Michael; Juel, Carsten

    2010-01-01

    , but is suggested primarily to participate in K+ release to the interstitium. Because there is restricted diffusion of K+ to the interstitium, K+ released to the T-tubules during AP propagation will be removed primarily by reuptake mediated by transport proteins located in the T-tubule membrane. The most important....... The relative content of the different K+-transporting proteins differs in oxidative and glycolytic muscles, and might explain the different [K+]e tolerance observed....

  13. Crystal structure of Δ(185-243)ApoA-I suggests a mechanistic framework for the protein adaptation to the changing lipid load in good cholesterol: from flatland to sphereland via double belt, belt buckle, double hairpin and trefoil/tetrafoil.

    Science.gov (United States)

    Gursky, Olga

    2013-01-09

    Apolipoprotein A-I (apoA-I) is the major protein of plasma high-density lipoproteins (HDLs), macromolecular assemblies of proteins and lipids that remove cell cholesterol and protect against atherosclerosis. HDL heterogeneity, large size (7.7-12 nm), and ability to exchange proteins have prevented high-resolution structural analysis. Low-resolution studies showed that two apoA-I molecules form an antiparallel α-helical "double belt" around an HDL particle. The atomic-resolution structure of the C-terminal truncated lipid-free Δ(185-243)apoA-I, determined recently by Mei and Atkinson, provides unprecedented new insights into HDL structure-function. It allows us to propose a molecular mechanism for the adaptation of the full-length protein to increasing lipid load during cholesterol transport. ApoA-I conformations on small, midsize, and large HDLs are proposed based on the tandem α-helical repeats and the crystal structure of Δ(185-243)apoA-I and are validated by comparison with extensive biophysical data reported by many groups. In our models, the central half of the double belt ("constant" segment 66-184) is structurally conserved while the N- and C-terminal half ("variable" segments 1-65 and 185-243) rearranges upon HDL growth. This includes incremental unhinging of the N-terminal bundle around two flexible regions containing G39 and G65 to elongate the belt, along with concerted swing motion of the double belt around G65-P66 and G185-G186 hinges that are aligned on various-size particles, to confer two-dimensional surface curvature to spherical HDLs. The proposed conformational ensemble integrates and improves several existing HDL models. It helps provide a structural framework necessary to understand functional interactions with over 60 other HDL-associated proteins and, ultimately, improve the cardioprotective function of HDL. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. Crystal Structure of Δ(185–243)apoA-I Suggests a Mechanistic Framework for the Protein Adaptation to the Changing Lipid Load in Good Cholesterol: From Flatland to Sphereland via Double Belt, Belt-Buckle, Double Hairpin and Trefoil/Tetrafoil

    Science.gov (United States)

    Gursky, Olga

    2012-01-01

    ApoA-I is the major protein of plasma high-density lipoproteins (HDL), macromolecular assemblies of proteins and lipids that remove cell cholesterol and protect against atherosclerosis. HDL heterogeneity, large size (7.7–12 nm) and ability to exchange proteins have prevented high-resolution structural analysis. Low-resolution studies showed that two apoA-I molecules form an antiparallel α-helical “double belt” around an HDL particle. Atomic-resolution structure of the C-terminal truncated lipid-free Δ(185–243)apoA-I, determined recently by Mei and Atkinson, provides unprecedented new insights into HDL structure-function. It allows us to propose a molecular mechanism for the adaptation of the full-length protein to increasing lipid load during cholesterol transport. ApoA-I conformations on the small, mid-size and large HDL are proposed based on the tandem α-helical repeats and the crystal structure of Δ(185–243)apoA-I, and are validated by comparison with extensive biophysical data reported by many groups. In our models, the central half of the double belt (“constant” segment 66–184) is structurally conserved while the N- and C-terminal half (“variable” segments 1–65 and 185–243) re-arranges upon HDL growth. This includes incremental unhinging of the N-terminal bundle around two flexible regions containing G39 and G65 to elongate the belt, along with concerted swing motion of the double belt around G65-P66 and G185–G186 hinges that are aligned on various-size particles, to confer 2D surface curvature to spherical HDL. The proposed conformational ensemble integrates and improves several existing HDL models. It helps provide a structural framework necessary to understand functional interactions with over 60 other HDL-associated proteins and, ultimately, improve cardioprotective function of HDL. PMID:23041415

  15. Computer-aided analyses of transport protein sequences: gleaning evidence concerning function, structure, biogenesis, and evolution.

    Science.gov (United States)

    Saier, M H

    1994-03-01

    Three-dimensional structures have been elucidated for very few integral membrane proteins. Computer methods can be used as guides for estimation of solute transport protein structure, function, biogenesis, and evolution. In this paper the application of currently available computer programs to over a dozen distinct families of transport proteins is reviewed. The reliability of sequence-based topological and localization analyses and the importance of sequence and residue conservation to structure and function are evaluated. Evidence concerning the nature and frequency of occurrence of domain shuffling, splicing, fusion, deletion, and duplication during evolution of specific transport protein families is also evaluated. Channel proteins are proposed to be functionally related to carriers. It is argued that energy coupling to transport was a late occurrence, superimposed on preexisting mechanisms of solute facilitation. It is shown that several transport protein families have evolved independently of each other, employing different routes, at different times in evolutionary history, to give topologically similar transmembrane protein complexes. The possible significance of this apparent topological convergence is discussed.

  16. Home-Use Tests - Cholesterol

    Science.gov (United States)

    ... Medical Procedures In Vitro Diagnostics Home Use Tests Cholesterol Share Tweet Linkedin Pin it More sharing options ... a home-use test kit to measure total cholesterol. What cholesterol is: Cholesterol is a fat (lipid) ...

  17. Herpes viral proteins manipulating the peptide transporter TAP

    NARCIS (Netherlands)

    Reits, E.; Griekspoor, A.; Neefjes, J.

    2002-01-01

    The peptide transporter associated with antigen processing (TAP) is crucial for class I-restricted antigen presentation because it transfers cytosolic peptides into the endoplasmic reticulum (ER) lumen for class I binding. It is therefore not surprising that TAP is targeted for inactivation by many

  18. Retrograde transport of protein toxins through the Golgi apparatus

    DEFF Research Database (Denmark)

    Sandvig, Kirsten; Skotland, Tore; van Deurs, Bo

    2013-01-01

    at the cell surface, and they are endocytosed both by clathrin-dependent and clathrin-independent mechanisms. Sorting to the Golgi and retrograde transport to the endoplasmic reticulum (ER) are common to these toxins, but the exact mechanisms turn out to be toxin and cell-type dependent. In the ER...

  19. Sex Differences in the Hepatic Cholesterol Sensing Mechanisms in Mice

    Directory of Open Access Journals (Sweden)

    Ingemar Björkhem

    2013-09-01

    Full Text Available Cholesterol is linked to many multifactorial disorders, including different forms of liver disease where development and severity depend on the sex. We performed a detailed analysis of cholesterol and bile acid synthesis pathways at the level of genes and metabolites combined with the expression studies of hepatic cholesterol uptake and transport in female and male mice fed with a high-fat diet with or without cholesterol. Lack of dietary cholesterol led to a stronger response of the sterol sensing mechanism in females, resulting in higher expression of cholesterogenic genes compared to males. With cholesterol in the diet, the genes were down-regulated in both sexes; however, males maintained a more efficient hepatic metabolic flux through the pathway. Females had higher content of hepatic cholesterol but this was likely not due to diminished excretion but rather due to increased synthesis and absorption. Dietary cholesterol and sex were not important for gallbladder bile acids composition. Neither sex up-regulated Cyp7a1 upon cholesterol loading and there was no compensatory up-regulation of Abcg5 or Abcg8 transporters. On the other hand, females had higher expression of the Ldlr and Cd36 genes. These findings explain sexual dimorphism of cholesterol metabolism in response to dietary cholesterol in a high-fat diet in mice, which contributes to understanding the sex-basis of cholesterol-associated liver diseases.

  20. Chemical activity of cholesterol in membranes.

    Science.gov (United States)

    Radhakrishnan, A; McConnell, H M

    2000-07-18

    Measurements are reported for the rate constants for the release of cholesterol (and dihydrocholesterol) to beta-cyclodextrin from mixtures with phospholipids in homogeneous monolayers at constant pressure at the air-water interface. In each mixture, it is found that the release rate shows a sharp decrease as the cholesterol concentration in the monolayer decreases through a composition corresponding to the stoichiometry of a cholesterol-phospholipid complex. The stoichiometry of the complex was established previously by the position of a sharp cusp in the thermodynamic phase diagram of each mixture and also by a minimum in average molecular area versus composition measurements. A theoretical model used earlier to account for the phase diagrams predicts the chemical potential and chemical activity of cholesterol in these mixtures. The calculated chemical activity also shows a sharp change at the complex stoichiometry in homogeneous monolayers. The similarities in change of observed release rate and calculated chemical activity are expected from reaction rate theory where the release rate is proportional to the cholesterol chemical activity. The chemical activity of cholesterol as determined by complex formation between some phospholipids and cholesterol in the plasma membrane of cells may serve a regulatory function with respect to intracellular cholesterol transport and biosynthesis.

  1. The actin cytoskeleton may control the polar distribution of an auxin transport protein

    Science.gov (United States)

    Muday, G. K.; Hu, S.; Brady, S. R.; Davies, E. (Principal Investigator)

    2000-01-01

    The gravitropic bending of plants has long been linked to the changes in the transport of the plant hormone auxin. To understand the mechanism by which gravity alters auxin movement, it is critical to know how polar auxin transport is initially established. In shoots, polar auxin transport is basipetal (i.e., from the shoot apex toward the base). It is driven by the basal localization of the auxin efflux carrier complex. One mechanism for localizing this efflux carrier complex to the basal membrane may be through attachment to the actin cytoskeleton. The efflux carrier protein complex is believed to consist of several polypeptides, including a regulatory subunit that binds auxin transport inhibitors, such as naphthylphthalamic acid (NPA). Several lines of experimentation have been used to determine if the NPA binding protein interacts with actin filaments. The NPA binding protein has been shown to partition with the actin cytoskeleton during detergent extraction. Agents that specifically alter the polymerization state of the actin cytoskeleton change the amount of NPA binding protein and actin recovered in these cytoskeletal pellets. Actin-affinity columns were prepared with polymers of actin purified from zucchini hypocotyl tissue. NPA binding activity was eluted in a single peak from the actin filament column. Cytochalasin D, which fragments the actin cytoskeleton, was shown to reduce polar auxin transport in zucchini hypocotyls. The interaction of the NPA binding protein with the actin cytoskeleton may localize it in one plane of the plasma membrane, and thereby control the polarity of auxin transport.

  2. Cholesterol-lowering effect of plant sterols.

    Science.gov (United States)

    AbuMweis, Suhad S; Jones, Peter J H

    2008-12-01

    Plant sterols are plant components that have a chemical structure similar to cholesterol except for the addition of an extra methyl or ethyl group; however, plant sterol absorption in humans is considerably less than that of cholesterol. In fact, plant sterols reduce cholesterol absorption and thus reduce circulating levels of cholesterol. Earlier studies that have tested the efficacy of plant sterols as cholesterol-lowering agents incorporated plant sterols into fat spreads. Later on, plant sterols were added to other food matrices, including juices, nonfat beverages, milk and yogurt, cheese, meat, croissants and muffins, and cereal and chocolate bars. The beneficial physiologic effects of plant sterols could be further enhanced by combining them with other beneficial substances, such as olive and fish oils, fibers, and soy proteins, or with exercise. The addition of plant sterols to the diet is suggested by health experts as a safe and effective way to reduce the risk of coronary heart disease.

  3. In vitro effects of selenite and mercuric chloride on liver thiobarbituric acid-reactive substances and non-protein thiols from rats: influences of dietary cholesterol and polyunsaturated and saturated fatty acids.

    Science.gov (United States)

    Farina, Marcelo; Soares, Félix Antunes; Feoli, Ana; Roehring, Cíntia; Brusque, Ana Maria; Rotta, Liane; Perry, Marcos Luis; Souza, Diogo Onofre; Rocha, João Batista T

    2003-06-01

    We measured the in vitro effects of mercuric chloride (Hg2+) and selenite (Se4+) on hepatic 2-thiobarbituric acid-reactive substances (TBARS) and non-protein sulfhydryl (NPSH) levels of rats fed diets enriched with polyunsaturated or saturated fatty acids with and without cholesterol. Male Wistar rats (21 d old) were assigned to one of four groups and fed diets containing 20% soybean oil, 20% soybean oil plus 1% cholesterol, 20% coconut oil, or coconut oil plus 1% cholesterol. After the feeding period (6 wk), body weight gain was equal in all groups. TBARS levels and NPSH content were measured after in vitro exposure to mercuric chloride (100 microM) and sodium selenite (25 microM) for 1 h. The lipid peroxidation, measured as TBARS levels in the control group, were statistically higher in hepatic homogenates of rats fed diets containing soybean oil than in groups fed coconut oil (P = 0.009). However, cholesterol supplementation did not change TBARS levels. Selenite alone did not modify TBARS production, whereas mercury alone significantly increased TBARS levels. Moreover, Se4+ protected against mercury-induced lipid peroxidation only in rats fed diets containing coconut oil. In the control group, dietary fat acids did not change NPSH levels. Selenite produced higher oxidative effects toward NPSH content, whereas Hg2+ decreased NPSH levels only in liver from rats fed diets containing soybean oil. NPSH levels were higher after concomitant exposure to Se4+ and Hg2+ chloride that after exposure to Se4+ alone, suggesting an interaction between Hg2+ and Se4+. Catalase activity was higher in animals fed diets containing soybean oil. Dietary cholesterol decreased glutathione peroxidase activity. Together these results indicated that the protective effect of Se4+ against mercury-induced lipid peroxidation depends on dietary fat saturation.

  4. ATPase and GTPase Tangos Drive Intracellular Protein Transport.

    Science.gov (United States)

    Shan, Shu-Ou

    2016-12-01

    The GTPase superfamily of proteins provides molecular switches to regulate numerous cellular processes. The 'GTPase switch' paradigm, in which external regulatory factors control the switch of a GTPase between 'on' and 'off' states, has been used to interpret the regulatory mechanism of many GTPases. However, recent work unveiled a class of nucleotide hydrolases that do not adhere to this classical paradigm. Instead, they use nucleotide-dependent dimerization cycles to regulate key cellular processes. In this review article, recent studies of dimeric GTPases and ATPases involved in intracellular protein targeting are summarized. It is suggested that these proteins can use the conformational plasticity at their dimer interface to generate multiple points of regulation, thereby providing the driving force and spatiotemporal coordination of complex cellular pathways. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Endocytic regulation of alkali metal transport proteins in mammals, yeast and plants.

    Science.gov (United States)

    Mulet, José Miguel; Llopis-Torregrosa, Vicent; Primo, Cecilia; Marqués, Ma Carmen; Yenush, Lynne

    2013-11-01

    The relative concentrations of ions and solutes inside cells are actively maintained by several classes of transport proteins, in many cases against their concentration gradient. These transport processes, which consume a large portion of cellular energy, must be constantly regulated. Many structurally distinct families of channels, carriers, and pumps have been characterized in considerable detail during the past decades and defects in the function of some of these proteins have been linked to a growing list of human diseases. The dynamic regulation of the transport proteins present at the cell surface is vital for both normal cellular function and for the successful adaptation to changing environments. The composition of proteins present at the cell surface is controlled on both the transcriptional and post-translational level. Post-translational regulation involves highly conserved mechanisms of phosphorylation- and ubiquitylation-dependent signal transduction routes used to modify the cohort of receptors and transport proteins present under any given circumstances. In this review, we will summarize what is currently known about one facet of this regulatory process: the endocytic regulation of alkali metal transport proteins. The physiological relevance, major contributors, parallels and missing pieces of the puzzle in mammals, yeast and plants will be discussed.

  6. Effects of isoflavone-containing soya protein on ex vivo cholesterol efflux, vascular function and blood markers of CVD risk in adults with moderately elevated blood pressure: a dose-response randomised controlled trial.

    Science.gov (United States)

    Richter, Chesney K; Skulas-Ray, Ann C; Fleming, Jennifer A; Link, Christina J; Mukherjea, Ratna; Krul, Elaine S; Kris-Etherton, Penny M

    2017-05-01

    Emerging CVD risk factors (e.g. HDL function and central haemodynamics) may account for residual CVD risk experienced by individuals who meet LDL-cholesterol and blood pressure (BP) targets. Recent evidence suggests that these emerging risk factors can be modified by polyphenol-rich interventions such as soya, but additional research is needed. This study was designed to investigate the effects of an isoflavone-containing soya protein isolate (delivering 25 and 50 g/d soya protein) on HDL function (i.e. ex vivo cholesterol efflux), macrovascular function and blood markers of CVD risk. Middle-aged adults (n 20; mean age=51·6 (sem 6·6) years) with moderately elevated brachial BP (mean systolic BP=129 (sem 9) mmHg; mean diastolic BP=82·5 (sem 8·4) mmHg) consumed 0 (control), 25 and 50 g/d soya protein in a randomised cross-over design. Soya and control powders were consumed for 6 weeks each with a 2-week compliance break between treatment periods. Blood samples and vascular function measures were obtained at baseline and following each supplementation period. Supplementation with 50 g/d soya protein significantly reduced brachial diastolic BP (-2·3 mmHg) compared with 25 g/d soya protein (Tukey-adjusted P=0·03) but not the control. Soya supplementation did not improve ex vivo cholesterol efflux, macrovascular function or other blood markers of CVD risk compared with the carbohydrate-matched control. Additional research is needed to clarify whether effects on these CVD risk factors depend on the relative health of participants and/or equol producing capacity.

  7. 9-Deazapurines as Broad-Spectrum Inhibitors of the ABC Transport Proteins P-Glycoprotein, Multidrug Resistance-Associated Protein 1, and Breast Cancer Resistance Protein.

    Science.gov (United States)

    Stefan, Katja; Schmitt, Sven Marcel; Wiese, Michael

    2017-11-09

    P-Glycoprotein (P-gp, ABCB1), multidrug resistance-associated protein 1 (MRP1, ABCC1), and breast cancer resistance protein (BCRP, ABCG2) are the three major ABC transport proteins conferring resistance to many structurally diverse anticancer agents, leading to the phenomenon called multidrug resistance (MDR). Much effort has been put into the development of clinically useful compounds to reverse MDR. Broad-spectrum inhibitors of ABC transport proteins can be of great use in cancers that simultaneously coexpress two or three transporters. In this work, we continued our effort to generate new, potent, nontoxic, and multiply effective inhibitors of the three major ABC transporters. The best compound was active in a very low micromolar concentration range against all three transporters and restored sensitivity toward daunorubicin (P-gp and MRP1) and SN-38 (BCRP) in A2780/ADR (P-gp), H69AR (MRP1), and MDCK II BCRP (BCRP) cells. Additionally, the compound is a noncompetitive inhibitor of daunorubicin (MRP1), calcein AM (P-gp), and pheophorbide A (BCRP) transport.

  8. The role of Monosaccharide Transport Proteins in carbohydrate assimilation, distribution, metabolism and homeostasis

    Science.gov (United States)

    Cura, Anthony J.; Carruthers, Anthony

    2012-01-01

    The facilitated diffusion of glucose, galactose, fructose, urate, myoinositol and dehydroascorbic acid in mammals is catalyzed by a family of 14 monosaccharide transport proteins called GLUTs. These transporters may be divided into 3 classes according to sequence similarity and function/substrate specificity. GLUT1 appears to be highly expressed in glycolytically active cells and has been co-opted in vitamin C auxotrophs to maintain the redox state of the blood through transport of dehydroascorbate. Several GLUTs are definitive glucose/galactose transporters, GLUT2 and GLUT5 are physiologically important fructose transporters, GLUT9 appears to be a urate transporter while GLUT13 (HMIT1) is a proton/myoinositol co-transporter. The physiologic substrates of some GLUTs remain to be established. The GLUTs are expressed in a tissue specific manner where affinity, specificity and capacity for substrate transport are paramount for tissue function. Although great strides have been made in characterizing GLUT-catalyzed monosaccharide transport and mapping GLUT membrane topography and determinants of substrate specificity, a unifying model for GLUT structure and function remains elusive. The GLUTs play a major role in carbohydrate homeostasis and the redistribution of sugar-derived carbons among the various organ systems. This is accomplished through a multiplicity of GLUT-dependent glucose sensing and effector mechanisms that regulate monosaccharide ingestion, absorption, distribution, cellular transport and metabolism and recovery/retention. Glucose transport and metabolism have co-evolved in mammals to support cerebral glucose utilization. PMID:22943001

  9. Choline transport via choline transporter-like protein 1 in conditionally immortalized rat syncytiotrophoblast cell lines TR-TBT.

    Science.gov (United States)

    Lee, N-Y; Choi, H-M; Kang, Y-S

    2009-04-01

    Choline is an essential nutrient for phospholipids and acetylcholine biosynthesis in normal development of fetus. In the present study, we investigated the functional characteristics of choline transport system and inhibitory effect of cationic drugs on choline transport in rat conditionally immortalized syncytiotrophoblast cell line (TR-TBT). Choline transport was weakly Na(+) dependent and significantly influenced by extracellular pH and by membrane depolarization. The transport process of choline is saturable with Michaelis-Menten constants (K(m)) of 68microM and 130microM in TR-TBT 18d-1 and TR-TBT 18d-2 respectively. Choline uptake in the cells was inhibited by unlabeled choline and hemicholinium-3 as well as various organic cations including guanidine, amiloride and acetylcholine. However, the prototypical organic cation tetraethylammonium and cimetidine showed very little inhibitory effect of choline uptake in TR-TBT cells. RT-PCR revealed that choline transporter-like protein 1 (CTL1) and organic cation transporter 2 (OCT2) are expressed in TR-TBT cells. The transport properties of choline in TR-TBT cells were similar or identical to that of CTL1 but not OCT2. CTL1 was also detected in human placenta. In addition, several cationic drugs such as diphenhydramine and verapamil competitively inhibited choline uptake in TR-TBT 18d-1 with K(i) of 115microM and 55microM, respectively. Our results suggest that choline transport system, which has intermediate affinity and weakly Na(+) dependent, in TR-TBT seems to occur through a CTL1 and this system may have relevance with the uptake of pharmacologically important organic cation drugs.

  10. Transport Vesicle Tethering at the Trans Golgi Network: Coiled Coil Proteins in Action.

    Science.gov (United States)

    Cheung, Pak-Yan P; Pfeffer, Suzanne R

    2016-01-01

    The Golgi complex is decorated with so-called Golgin proteins that share a common feature: a large proportion of their amino acid sequences are predicted to form coiled-coil structures. The possible presence of extensive coiled coils implies that these proteins are highly elongated molecules that can extend a significant distance from the Golgi surface. This property would help them to capture or trap inbound transport vesicles and to tether Golgi mini-stacks together. This review will summarize our current understanding of coiled coil tethers that are needed for the receipt of transport vesicles at the trans Golgi network (TGN). How do long tethering proteins actually catch vesicles? Golgi-associated, coiled coil tethers contain numerous binding sites for small GTPases, SNARE proteins, and vesicle coat proteins. How are these interactions coordinated and are any or all of them important for the tethering process? Progress toward understanding these questions and remaining, unresolved mysteries will be discussed.

  11. Role of STARD4 in sterol transport between the endocytic recycling compartment and the plasma membrane.

    Science.gov (United States)

    Iaea, David B; Mao, Shu; Lund, Frederik W; Maxfield, Frederick R

    2017-04-15

    Cholesterol is an essential constituent of membranes in mammalian cells. The plasma membrane and the endocytic recycling compartment (ERC) are both highly enriched in cholesterol. The abundance and distribution of cholesterol among organelles are tightly controlled by a combination of mechanisms involving vesicular and nonvesicular sterol transport processes. Using the fluorescent cholesterol analogue dehydroergosterol, we examined sterol transport between the plasma membrane and the ERC using fluorescence recovery after photobleaching and a novel sterol efflux assay. We found that sterol transport between these organelles in a U2OS cell line has a t 1/2 =12-15 min. Approximately 70% of sterol transport is ATP independent and therefore is nonvesicular. Increasing cellular cholesterol levels dramatically increases bidirectional transport rate constants, but decreases in cholesterol levels have only a modest effect. A soluble sterol transport protein, STARD4, accounts for ∼25% of total sterol transport and ∼33% of nonvesicular sterol transport between the plasma membrane and ERC. This study shows that nonvesicular sterol transport mechanisms and STARD4 in particular account for a large fraction of sterol transport between the plasma membrane and the ERC. © 2017 Iaea et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  12. Effects of apolipoproteins on the kinetics of cholesterol exchange

    International Nuclear Information System (INIS)

    Letizia, J.Y.; Phillips, M.C.

    1991-01-01

    The effects of apolipoproteins on the kinetics of cholesterol exchange have been investigated by monitoring the transfer of [ 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 [ 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 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

  13. Carcinoembryonic antigen-related cell adhesion molecule 1 is the 85-kilodalton pronase-resistant biliary gycoprotein in the cholesterol crystallization promoting low density protein-lipid complex

    Czech Academy of Sciences Publication Activity Database

    Jirsa, M.; Muchová, L.; Dráberová, Lubica; Dráber, Petr; Šmíd, F.; Kuroki, M.; Mareček, Z.; Groen, A. K.

    2001-01-01

    Roč. 34, - (2001), s. 1075-1082 ISSN 0270-9139 R&D Projects: GA MŠk LN00A026; GA MZd IZ4046 Keywords : carcinoembryonic antigen * biliary glycoprotein * cholesterol Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.096, year: 2001

  14. Circulating levels of linoleic acid and HDL-cholesterol are major determinants of 4-hydroxynonenal protein adducts in patients with heart failure

    Directory of Open Access Journals (Sweden)

    Caroline Asselin

    2014-01-01

    Conclusion: Results from this study emphasize the importance of considering changes in lipids and lipoproteins in the interpretation of measurements of lipid peroxidation products. Further studies appear warranted to explore the possibility that HDL-cholesterol particles may be a carrier of 4HNE adducts.

  15. The human synaptic vesicle protein, SV2A, functions as a galactose transporter in Saccharomyces cerevisiae.

    Science.gov (United States)

    Madeo, Marianna; Kovács, Attila D; Pearce, David A

    2014-11-28

    SV2A is a synaptic vesicle membrane protein expressed in neurons and endocrine cells and involved in the regulation of neurotransmitter release. Although the exact function of SV2A still remains elusive, it was identified as the specific binding site for levetiracetam, a second generation antiepileptic drug. Our sequence analysis demonstrates that SV2A has significant homology with several yeast transport proteins belonging to the major facilitator superfamily (MFS). Many of these transporters are involved in sugar transport into yeast cells. Here we present evidence showing, for the first time, that SV2A is a galactose transporter. We expressed human SV2A in hexose transport-deficient EBY.VW4000 yeast cells and demonstrated that these cells are able to grow on galactose-containing medium but not on other fermentable carbon sources. Furthermore, the addition of the SV2A-binding antiepileptic drug levetiracetam to the medium inhibited the galactose-dependent growth of hexose transport-deficient EBY.VW4000 yeast cells expressing human SV2A. Most importantly, direct measurement of galactose uptake in the same strain verified that SV2A is able to transport extracellular galactose inside the cells. The newly identified galactose transport capability of SV2A may have an important role in regulating/modulating synaptic function. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Plasma membrane microdomains regulate turnover of transport proteins in yeast

    Czech Academy of Sciences Publication Activity Database

    Grossmann, G.; Malínský, Jan; Stahlschmidt, W.; Loibl, M.; Weig-Meckl, I.; Frommer, W.B.; Opekarová, Miroslava; Tanner, W.

    2008-01-01

    Roč. 183, č. 6 (2008), s. 1075-1088 ISSN 0021-9525 R&D Projects: GA ČR GA204/06/0009; GA ČR GA204/07/0133; GA ČR GC204/08/J024 Institutional research plan: CEZ:AV0Z50390703; CEZ:AV0Z50200510 Keywords : Lithium acetate * Membrane compartment of Can1 * Monomeric red fluorescent protein Subject RIV: EA - Cell Biology Impact factor: 9.120, year: 2008

  17. Resveratrol Inhibits Porcine Intestinal Glucose and Alanine Transport: Potential Roles of Na+/K+-ATPase Activity, Protein Kinase A, AMP-Activated Protein Kinase and the Association of Selected Nutrient Transport Proteins with Detergent Resistant Membranes

    Directory of Open Access Journals (Sweden)

    Stefanie Klinger

    2018-03-01

    Full Text Available Background: Beneficial effects of Resveratrol (RSV have been demonstrated, including effects on transporters and channels. However, little is known about how RSV influences intestinal transport. The aim of this study was to further characterize the effects of RSV on intestinal transport and the respective mechanisms. Methods: Porcine jejunum and ileum were incubated with RSV (300 µM, 30 min in Ussing chambers (functional studies and tissue bathes (detection of protein expression, phosphorylation, association with detergent resistant membranes (DRMs. Results: RSV reduced alanine and glucose-induced short circuit currents (ΔIsc and influenced forskolin-induced ΔIsc. The phosphorylation of sodium–glucose-linked transporter 1 (SGLT1, AMP-activated protein kinase (AMPK, protein kinase A substrates (PKA-S and liver kinase B1 (LKB1 increased but a causative relation to the inhibitory effects could not directly be established. The DRM association of SGLT1, peptide transporter 1 (PEPT1 and (phosphorylated Na+/H+-exchanger 3 (NHE3 did not change. Conclusion: RSV influences the intestinal transport of glucose, alanine and chloride and is likely to affect other transport processes. As the effects of protein kinase activation vary between the intestinal localizations, it would appear that increasing cyclic adenosine monophosphate (cAMP levels are part of the mechanism. Nonetheless, the physiological responses depend on cell type-specific structures.

  18. Effect of physical training on glucose transporter protein and mRNA levels in rat adipocytes

    DEFF Research Database (Denmark)

    Stallknecht, B; Andersen, P H; Vinten, J

    1993-01-01

    Physical training increases insulin-stimulated glucose transport and the number of glucose transporters in adipocytes measured by cytochalasin B binding. In the present study we used immunoblotting to measure the abundance of two glucose transporters (GLUT-4, GLUT-1) in white adipocytes from...... trained rats. Furthermore, the abundance of the mRNAs for these proteins and glucose transport was measured. Rats were swim-trained for 10 wk, and adipocytes were isolated from epididymal fat pads. The amount of GLUT-4/adipocyte volume unit was significantly higher in trained animals compared with both...... age- and cell size-matched animals. The amount of GLUT-4 mRNA was also increased by training and it decreased with increasing age. Furthermore, young age as well as training was accompanied by relatively low GLUT-4 protein/mRNA and relatively high overall GLUT-4 efficiency (recruitability and...

  19. Major intrinsic proteins and arsenic transport in plants: new players and their potential role.

    Science.gov (United States)

    Bienert, Gerd P; Jahn, Thomas P

    2010-01-01

    Arsenic (As) is a toxic and highly abundant metalloid that endangers human health through drinking water and the food chain. The most common forms of As in the environment re arsenate [As(V)] and arsenite [As(III)]. As(V) is a nonfunctional phosphate analog that enters the food chain via plant phosphate transporters. Recently, evidence was provided that uptake of As(III)--the second most abundant As species in soils--is mediated by plant nodulin26-like intrinsic proteins (NIPs), a subfamily of plant major intrinsic proteins (MIPs). Specific NIPs are also essential for the uptake of the metalloids boron and silicon and aquaglyceroporins from microbes and mammals were shown to be the major routes of As uptake. Therefore As(III) transport through MIPs is a conserved and ancient feature. In this chapter we summarize the current view on As transport in plants and address the potential physiological significance of As(III) transport through NIPs.

  20. Phloem RNA-binding proteins as potential components of the long-distance RNA transport system.

    Directory of Open Access Journals (Sweden)

    VICENTE ePALLAS

    2013-05-01

    Full Text Available RNA-binding proteins (RBPs govern a myriad of different essential processes in eukaryotic cells. Recent evidence reveals that apart from playing critical roles in RNA metabolism and RNA transport, RBPs perform a key function in plant adaption to various environmental conditions. Long distance RNA transport occurs in land plants through the phloem, a conducting tissue that integrates the wide range of signalling pathways required to regulate plant development and response to stress processes. The macromolecules in the phloem pathway vary greatly and include defence proteins, transcription factors, chaperones acting in long distance trafficking, and RNAs (mRNAs, siRNAs and miRNAs. How these RNA molecules translocate through the phloem is not well understood, but recent evidence indicates the presence of translocatable RNA-binding proteins in the phloem, which act as potential components of long distance RNA transport system. This review updates our knowledge on the characteristics and functions of RBPs present in the phloem.

  1. Comparative genomic analyses of transport proteins encoded within the genomes of Leptospira species.

    Science.gov (United States)

    Buyuktimkin, Bora; Saier, Milton H

    2015-11-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, Leptospira interrogans serovar Lai str. 56601 and Leptospira borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, Leptospira biflexa serovar Patoc str. 'Patoc 1 (Ames)'. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity was accompanied by progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Comparative analyses of transport proteins encoded within the genomes of Leptospira species.

    Science.gov (United States)

    Buyuktimkin, Bora; Saier, Milton H

    2016-09-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, Leptospira interrogans serovar Lai str. 56601 and Leptospira borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, Leptospira biflexa serovar Patoc str. 'Patoc 1 (Ames)'. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they all have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity arose in Leptospira correlating to progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. Copyright © 2016. Published by Elsevier Ltd.

  3. Interaction between the glutamate transporter GLT1b and the synaptic PDZ domain protein PICK1

    DEFF Research Database (Denmark)

    Bassan, Merav; Liu, Hongguang; Madsen, Kenneth L

    2008-01-01

    Synaptic plasticity is implemented by the interaction of glutamate receptors with PDZ domain proteins. Glutamate transporters provide the only known mechanism of clearance of glutamate from excitatory synapses, and GLT1 is the major glutamate transporter. We show here that GLT1 interacts...... expressing PICK1 and GLT1b. In addition, expression of GLT1b in COS7 cells changed the distribution of PICK1, bringing it to the surface. GLT1b and PICK1 co-localized with each other and with synaptic markers in hippocampal neurons in culture. Phorbol ester, an activator of protein kinase C (PKC), a known...

  4. Frog oocytes to unveil the structure and supramolecular organization of human transport proteins.

    Directory of Open Access Journals (Sweden)

    Marc J Bergeron

    Full Text Available Structural analyses of heterologously expressed mammalian membrane proteins remain a great challenge given that microgram to milligram amounts of correctly folded and highly purified proteins are required. Here, we present a novel method for the expression and affinity purification of recombinant mammalian and in particular human transport proteins in Xenopus laevis frog oocytes. The method was validated for four human and one murine transporter. Negative stain transmission electron microscopy (TEM and single particle analysis (SPA of two of these transporters, i.e., the potassium-chloride cotransporter 4 (KCC4 and the aquaporin-1 (AQP1 water channel, revealed the expected quaternary structures within homogeneous preparations, and thus correct protein folding and assembly. This is the first time a cation-chloride cotransporter (SLC12 family member is isolated, and its shape, dimensions, low-resolution structure and oligomeric state determined by TEM, i.e., by a direct method. Finally, we were able to grow 2D crystals of human AQP1. The ability of AQP1 to crystallize was a strong indicator for the structural integrity of the purified recombinant protein. This approach will open the way for the structure determination of many human membrane transporters taking full advantage of the Xenopus laevis oocyte expression system that generally yields robust functional expression.

  5. Insulin-like growth factor binding protein 3 has opposing actions on malignant and nonmalignant breast epithelial cells that are each reversible and dependent upon cholesterol-stabilized integrin receptor complexes.

    Science.gov (United States)

    Burrows, C; Holly, J M P; Laurence, N J; Vernon, E G; Carter, J V; Clark, M A; McIntosh, J; McCaig, C; Winters, Z E; Perks, C M

    2006-07-01

    IGF-binding protein (IGFBP)-3 is generally considered to have actions that counterbalance those of IGFs and is therefore being developed as a cancer treatment. In breast tumors, however, high levels are associated with aggressive tumors and poor prognosis. Consistent with this we have demonstrated that although IGFBP-3 and a non-IGF-binding fragment (serine phosphorylation domain peptide) reduced attachment and enhanced apoptosis of Hs578T breast cancer cells cultured on collagen or laminin, it promoted their attachment and survival on fibronectin, which is abundant in the matrix of aggressive tumors. We have now examined the factors that determine whether IGFBP-3 has positive or negative actions on breast epithelial cells. IGFBP-3 also promoted survival of Hs578T cells in the presence of an antibody to the beta1-integrin subunit or when cholesterol-stabilized complexes were disrupted. These actions were blocked by IGF-I or a MAPK inhibitor. Serine phosphorylation domain peptide had similar actions on MCF-7 cells that were again reversed on fibronectin or with disruption of cholesterol-stabilized complexes and blocked by the beta1-integrin antibody. In contrast, IGFBP-3 promoted growth and survival for nonmalignant MCF-10A cells, but these effects were again reversed on fibronectin and blocked by the beta1 antibody or a MAPK inhibitor or by disruption of cholesterol-stabilized complexes. On Hs578T cells, IGFBP-3 bound to caveolin-1 and beta1-integrins, enhancing their aggregation, the recruitment of focal adhesion kinase, and the activation of MAPK. In summary, with three breast epithelial cell lines, IGFBP-3 had positive or negative effects on growth and survival dependent upon the status of cholesterol-stabilized integrin receptor complexes.

  6. Cholesterol - what to ask your doctor

    Science.gov (United States)

    ... your doctor; What to ask your doctor about cholesterol ... What is my cholesterol level? What should my cholesterol level be? What are HDL ("good") cholesterol and LDL ("bad") cholesterol? Does my cholesterol ...

  7. National Cholesterol Education Month

    Centers for Disease Control (CDC) Podcasts

    2009-09-01

    Do you know your cholesterol numbers? Your doctor can do a simple test to check your cholesterol levels and help you make choices that lower your risk for heart disease and stroke.  Created: 9/1/2009 by National Center for Chronic Disease Prevention and Health Promotion (NCCDPHP).   Date Released: 9/9/2009.

  8. Cholesterol and Health

    Indian Academy of Sciences (India)

    catalysed reactions. Keywords. Cholesterol,levelofcholesterol, dietary regimen, LDl, HDl. Figure 1. Structure of cho- lesterol. Steroids occur widely in both plants and animals; the important steroids however, are found in animals where they have various essential biological functions. The most abundant steroid is cholesterol.

  9. RECOVERY ACT - Thylakoid Assembly and Folded Protein Transport by the Tat Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Dabney-Smith, Carole [Miami Univ., Oxford, OH (United States)

    2016-07-18

    Assembly of functional photosystems complete with necessary intrinsic (membrane-bound) and extrinsic proteins requires the function of at least 3 protein transport pathways in thylakoid membranes. Our research focuses on one of those pathways, a unique and essential protein transport pathway found in the chloroplasts of plants, bacteria, and some archaebacteria, the Twin arginine translocation (Tat) system. The chloroplast Tat (cpTat) system is thought to be responsible for the proper location of ~50% of thylakoid lumen proteins, several of which are necessary for proper photosystem assembly, maintenance, and function. Specifically, cpTat systems are unique because they transport fully folded and assembled proteins across ion tight membranes using only three membrane components, Tha4, Hcf106, and cpTatC, and the protonmotive force generated by photosynthesis. Despite the importance of the cpTat system in plants, the mechanism of transport of a folded precursor is not well known. Our long-term goal is to investigate the role protein transport systems have on organelle biogenesis, particularly the assembly of membrane protein complexes in thylakoids of chloroplasts. The objective of this proposal is to correlate structural changes in the membrane-bound cpTat component, Tha4, to the mechanism of translocation of folded-precursor substrates across the membrane bilayer by using a cysteine accessibility and crosslinking approach. Our central hypothesis is that the precursor passes through a proteinaceous pore of assembled Tha4 protomers that have undergone a conformational or topological change in response to transport. This research is predicated upon the observations that Tha4 exists in molar excess in the membrane relative to the other cpTat components; its regulated assembly to the precursor-bound receptor; and our data showing oligomerization of Tha4 into very large complexes in response to transport. Our rationale for these studies is that understanding cp

  10. Phosphatidylcholine: cholesterol phase diagrams.

    Science.gov (United States)

    Thewalt, J L; Bloom, M

    1992-10-01

    Two mono-cis-unsaturated phosphatidylcholine (PC) lipid molecules, having very different gel-liquid crystalline phase transition temperatures as a consequence of the relative positions of the double bond, exhibit PC:cholesterol phase diagrams that are very similar to each other and to that obtained previously for a fully saturated PC:cholesterol mixture (Vist, M. R., and J. H. Davis. 1990. Biochemistry 29:451-464). This leads to the conjecture that PC:cholesterol membrane phase diagrams have a universal form which is relatively independent of the precise chemical structure of the PC molecule. One feature of this phase diagram is the observation over a wide temperature range of a fluid but highly conformationally ordered phase at bilayer concentrations of more than approximately 25 mol% cholesterol. This ;liquid ordered' phase is postulated to be the relevant physical state for many biological membranes, such as the plasma membrane of eukaryotic cells, that contain substantial amounts of cholesterol or equivalent sterols.

  11. Regorafenib is transported by the organic anion transporter 1B1 and the multidrug resistance protein 2.

    Science.gov (United States)

    Ohya, Hiroki; Shibayama, Yoshihiko; Ogura, Jiro; Narumi, Katsuya; Kobayashi, Masaki; Iseki, Ken

    2015-01-01

    Regorafenib is a small molecule inhibitor of tyrosine kinases, and has been shown to improve the outcomes of patients with advanced colorectal cancer and advanced gastrointestinal stromal tumors. The transport profiles of regorafenib by various transporters were evaluated. HEK293/organic anion transporting polypeptide 1B1 (OATP1B1) cells exhibited increased drug sensitivity to regorafenib. Regorafenib inhibited the uptake of 3H-estrone sulfate by HEK293/OATP1B1 cells in a dose-dependent manner, but did not affect its elimination by P-glycoproteins. The concentration of regorafenib was significantly lower in LLC-PK1/multidrug resistance protein 2 (MRP2) cells than in LLC-PK1 cells treated with the MRP2 inhibitor, MK571. MK571 abolished the inhibitory effects of regorafenib on intracellular accumulation in LLC-PK1/MRP2 cells. The uptake of regorafenib was significantly higher in HEK293/OATP1B1 cells than in OATP1B1-mock cells. Transport kinetics values were estimated to be Km=15.9 µM and Vmax=1.24 nmol/mg/min. No significant difference was observed in regorafenib concentrations between HEK293/OATP1B3 and OATP1B3-mock cells. These results indicated that regorafenib is a substrate for MRP2 and OATP1B1, and also suggest that the substrate preference of regorafenib may implicate the pharmacokinetic profiles of regorafenib.

  12. Electronic transport on the spatial structure of the protein: Three-dimensional lattice model

    International Nuclear Information System (INIS)

    Sarmento, R.G.; Frazão, N.F.; Macedo-Filho, A.

    2017-01-01

    Highlights: • The electronic transport on the structure of the three-dimensional lattice model of the protein is studied. • The signing of the current–voltage is directly affected by permutations of the weak bonds in the structure. • Semiconductor behave of the proteins suggest a potential application in the development of novel biosensors. - Abstract: We report a numerical analysis of the electronic transport in protein chain consisting of thirty-six standard amino acids. The protein chains studied have three-dimensional structure, which can present itself in three distinct conformations and the difference consist in the presence or absence of thirteen hydrogen-bondings. Our theoretical method uses an electronic tight-binding Hamiltonian model, appropriate to describe the protein segments modeled by the amino acid chain. We note that the presence and the permutations between weak bonds in the structure of proteins are directly related to the signing of the current–voltage. Furthermore, the electronic transport depends on the effect of temperature. In addition, we have found a semiconductor behave in the models investigated and it suggest a potential application in the development of novel biosensors for molecular diagnostics.

  13. Electronic transport on the spatial structure of the protein: Three-dimensional lattice model

    Energy Technology Data Exchange (ETDEWEB)

    Sarmento, R.G. [Departamento de Ciências Biológicas, Universidade Federal do Piauí, 64800-000 Floriano, PI (Brazil); Frazão, N.F. [Centro de Educação e Saúde, Universidade Federal de Campina Grande, 581750-000 Cuité, PB (Brazil); Macedo-Filho, A., E-mail: amfilho@gmail.com [Campus Prof. Antonio Geovanne Alves de Sousa, Universidade Estadual do Piauí, 64260-000 Piripiri, PI (Brazil)

    2017-01-30

    Highlights: • The electronic transport on the structure of the three-dimensional lattice model of the protein is studied. • The signing of the current–voltage is directly affected by permutations of the weak bonds in the structure. • Semiconductor behave of the proteins suggest a potential application in the development of novel biosensors. - Abstract: We report a numerical analysis of the electronic transport in protein chain consisting of thirty-six standard amino acids. The protein chains studied have three-dimensional structure, which can present itself in three distinct conformations and the difference consist in the presence or absence of thirteen hydrogen-bondings. Our theoretical method uses an electronic tight-binding Hamiltonian model, appropriate to describe the protein segments modeled by the amino acid chain. We note that the presence and the permutations between weak bonds in the structure of proteins are directly related to the signing of the current–voltage. Furthermore, the electronic transport depends on the effect of temperature. In addition, we have found a semiconductor behave in the models investigated and it suggest a potential application in the development of novel biosensors for molecular diagnostics.

  14. Structure-Functional Basis of Ion Transport in Sodium–Calcium Exchanger (NCX Proteins

    Directory of Open Access Journals (Sweden)

    Moshe Giladi

    2016-11-01

    Full Text Available The membrane-bound sodium–calcium exchanger (NCX proteins shape Ca2+ homeostasis in many cell types, thus participating in a wide range of physiological and pathological processes. Determination of the crystal structure of an archaeal NCX (NCX_Mj paved the way for a thorough and systematic investigation of ion transport mechanisms in NCX proteins. Here, we review the data gathered from the X-ray crystallography, molecular dynamics simulations, hydrogen–deuterium exchange mass-spectrometry (HDX-MS, and ion-flux analyses of mutants. Strikingly, the apo NCX_Mj protein exhibits characteristic patterns in the local backbone dynamics at particular helix segments, thereby possessing characteristic HDX profiles, suggesting structure-dynamic preorganization (geometric arrangements of catalytic residues before the transition state of conserved α1 and α2 repeats at ion-coordinating residues involved in transport activities. Moreover, dynamic preorganization of local structural entities in the apo protein predefines the status of ion-occlusion and transition states, even though Na+ or Ca2+ binding modifies the preceding backbone dynamics nearby functionally important residues. Future challenges include resolving the structural-dynamic determinants governing the ion selectivity, functional asymmetry and ion-induced alternating access. Taking into account the structural similarities of NCX_Mj with the other proteins belonging to the Ca2+/cation exchanger superfamily, the recent findings can significantly improve our understanding of ion transport mechanisms in NCX and similar proteins.

  15. Integrated Translatomics with Proteomics to Identify Novel Iron–Transporting Proteins in Streptococcus pneumoniae

    Directory of Open Access Journals (Sweden)

    Xiao-Yan eYang

    2016-02-01

    Full Text Available Streptococcus pneumoniae (S. pneumoniae is a major human pathogen causing morbidity and mortality worldwide. Efficiently acquiring iron from the environment is critical for S. pneumoniae to sustain growth and cause infection. There are only three known iron-uptake systems in Streptococcal species responsible for iron acquisition from the host, including ABC transporters PiaABC, PiuABC and PitABC. Besides, no other iron-transporting system has been suggested. In this work, we employed our newly established translating mRNA analysis integrated with proteomics to evaluate the possible existence of novel iron transporters in the bacterium. We simultaneously deleted the iron-binding protein genes of the three iron-uptake systems to construct a piaA/piuA/pitA triple mutant (Tri-Mut of S. pneumoniae D39, in which genes and proteins related to iron transport should be regulated in response to the deletion. With ribosome associated mRNA sequencing-based translatomics focusing on translating mRNA and iTRAQ quantitative proteomics based on the covalent labeling of peptides with tags of varying mass, we indeed observed a large number of genes and proteins representing various coordinated biological pathways with significantly altered expression levels in the Tri-Mut mutant. Highlighted in this observation is the identification of several new potential iron-uptake ABC transporters participating in iron metabolism of Streptococcus. In particular, putative protein SPD_1609 in operon 804 was verified to be a novel iron-binding protein with similar function to PitA in S. pneumoniae. These data derived from the integrative translatomics and proteomics analyses provided rich information and insightful clues for further investigations on iron-transporting mechanism in bacteria and the interplay between Streptococcal iron availability and the biological metabolic pathways.

  16. Cysteine-rich intestinal protein binds zinc during transmucosal zinc transport

    International Nuclear Information System (INIS)

    Hempe, J.M.; Cousins, R.J.

    1991-01-01

    The mechanism of zinc absorption has not been delineated, but kinetic studies show that both passive and carrier-mediated processes are involved. The authors have identified a low molecular mass zinc-binding protein in the soluble fraction of rat intestinal mucosa that could function as an intracellular zinc carrier. The protein was not detected in liver or pancreas, suggesting a role specific to the intestine. The protein binds zinc during transmucosal zinc transport and shows signs of saturation at higher luminal zinc concentrations, characteristics consistent with a role in carrier-mediated zinc absorption. Microsequence analysis of the protein purified by gel-filtration HPCL and SDS/PAGE showed complete identity within the first 41 N-terminal amino acids with the deduced protein sequence of cysteine-rich intestinal protein. These investigators showed that the gene for this protein is developmentally regulated in neonates during the suckling period, conserved in many vertebrate species, and predominantly expressed in the small intestine. Cysteine-rich intestinal protein contains a recently identified conserved sequence of histidine and cysteine residues, the LIM motif, which our results suggest confers metal-binding properties that are important for zinc transport and/or functions of this micronutrient

  17. Physiological and pathological implications of cholesterol.

    Science.gov (United States)

    Cortes, Victor A; Busso, Dolores; Maiz, Alberto; Arteaga, Antonio; Nervi, Flavio; Rigotti, Attilio

    2014-01-01

    Cholesterol has evolved to fulfill sophisticated biophysical, cell signaling and endocrine requirements of animal systems. At a cellular level, cholesterol is found in membranes, where it increases both bilayer stiffness and impermeability to water and ions. Furthermore, cholesterol is integrated into specialized lipid-protein membrane microdomains with critical topographical and signaling functions. At an organismal level, cholesterol is the precursor for all steroid hormones, including gluco- and mineralo-corticoids, sex hormones and vitamin D, all of which regulate carbohydrate, sodium, reproductive and bone homeostasis, respectively. This sterol is also the precursor for bile acids, which are important for intestinal absorption of dietary lipids as well as energy and glucose metabolic regulation. Importantly, complex mechanisms maintain cholesterol within physiological ranges and the disregulation of these mechanisms results in embryonic or adult diseases, caused by either excessive or reduced tissue cholesterol levels. The causative role of cholesterol in these diseases has been demonstrated by diverse genetic and pharmacologic animal models that are commented in this review.

  18. A Closer Look at Cardioprotective Function of HDL: Revise the HDL – Cholesterol Hypothesis?

    Directory of Open Access Journals (Sweden)

    Anna Meiliana

    2014-04-01

    Full Text Available BACKGROUND: The strong inverse association of plasma levels of high-density lipoprotein (HDL cholesterol with coronary heart disease (CHD found in human epidemiological studies led to the development of the ‘HDL cholesterol hypothesis’, which posits that intervention to raise HDL cholesterol will result in reduced risk of CHD. A number of recent developments have brought the potential protective role of HDL into question. Several clinical trials of agents that substantially raise HDL-C have been demonstrated to not reduce CHD event rates. CONTENT: For decades, HDL and HDL-cholesterol (HDL-C levels were viewed as synonymous, and modulation of HDL-C levels by drug therapy held great promise for the prevention and treatment of cardiovascular disease. Nevertheless, recent failures of drugs that raise HDL-C to reduce cardiovascular risk and the now greater understanding of the complexity of HDL composition and biology have prompted researchers in the field to redefine HDL. As such, the focus of HDL has now started to shift away from a cholesterol-centric view toward HDL particle number, subclasses, and other alternative metrics of HDL. Many of the recently discovered functions of HDL are, in fact, not strictly conferred by its ability to promote cholesterol flux but by the other molecules it transports, including a diverse set of proteins, small RNAs, hormones, carotenoids, vitamins, and bioactive lipids. Based on HDL’s ability to interact with almost all cells and deliver fat-soluble cargo, HDL has the remarkable capacity to affect a wide variety of endocrine-like systems. SUMMARY: There is a significant need to redefine HDL and its benefit. HDL transports a diverse set of functional proteins, including many binding proteins. HDL transports and deliver vitamins, carotenoids, and other small molecules. Moreover, HDL transports hormones, steroids and bile acids, and can modulate multiple endocrine pathways. HDLs also transport and deliver micro

  19. Inhibiting Cholesterol Absorption During Lactation Programs Future Intestinal Absorption of Cholesterol in Adult Mice.

    Science.gov (United States)

    Dimova, Lidiya G; de Boer, Jan Freark; Plantinga, Josee; Plösch, Torsten; Hoekstra, Menno; Verkade, Henkjan J; Tietge, Uwe J F

    2017-08-01

    In nematodes, the intestine senses and integrates early life dietary cues that lead to lifelong epigenetic adaptations to a perceived nutritional environment-it is not clear whether this process occurs in mammals. We aimed to establish a mouse model of reduced dietary cholesterol availability from maternal milk and investigate the consequences of decreased milk cholesterol availability, early in life, on the metabolism of cholesterol in adult mice. We blocked intestinal absorption of cholesterol in milk fed to newborn mice by supplementing the food of dams (for 3 weeks between birth and weaning) with ezetimibe, which is secreted into milk. Ezetimibe interacts with the intestinal cholesterol absorption transporter NPC1l1 to block cholesterol uptake into enterocytes. Characterization of these offspring at 24 weeks of age showed a 27% decrease in cholesterol absorption (P intestine. We observed increased histone H3K9me3 methylation at positions -423 to -607 of the proximal Npc1l1 promoter in small intestine tissues from 24-week-old offspring fed ezetimibe during lactation, compared with controls. These findings show that the early postnatal mammalian intestine functions as an environmental sensor of nutritional conditions, responding to conditions such as low cholesterol levels by epigenetic modifications of genes. Further studies are needed to determine how decreased sterol absorption for a defined period might activate epigenetic regulators; the findings of our study might have implications for human infant nutrition and understanding and preventing cardiometabolic disease. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

  20. Inhibition of retrograde transport modulates misfolded protein accumulation and clearance in motoneuron diseases.

    Science.gov (United States)

    Cristofani, Riccardo; Crippa, Valeria; Rusmini, Paola; Cicardi, Maria Elena; Meroni, Marco; Licata, Nausicaa V; Sala, Gessica; Giorgetti, Elisa; Grunseich, Christopher; Galbiati, Mariarita; Piccolella, Margherita; Messi, Elio; Ferrarese, Carlo; Carra, Serena; Poletti, Angelo

    2017-08-03

    Motoneuron diseases, like spinal bulbar muscular atrophy (SBMA) and amyotrophic lateral sclerosis (ALS), are associated with proteins that because of gene mutation or peculiar structures, acquire aberrant (misfolded) conformations toxic to cells. To prevent misfolded protein toxicity, cells activate a protein quality control (PQC) system composed of chaperones and degradative pathways (proteasome and autophagy). Inefficient activation of the PQC system results in misfolded protein accumulation that ultimately leads to neuronal cell death, while efficient macroautophagy/autophagy-mediated degradation of aggregating proteins is beneficial. The latter relies on an active retrograde transport, mediated by dynein and specific chaperones, such as the HSPB8-BAG3-HSPA8 complex. Here, using cellular models expressing aggregate-prone proteins involved in SBMA and ALS, we demonstrate that inhibition of dynein-mediated retrograde transport, which impairs the targeting to autophagy of misfolded species, does not increase their aggregation. Rather, dynein inhibition correlates with a reduced accumulation and an increased clearance of mutant ARpolyQ, SOD1, truncated TARDBP/TDP-43 and expanded polyGP C9ORF72 products. The enhanced misfolded protein clearance is mediated by the proteasome, rather than by autophagy and correlates with the upregulation of the HSPA8 cochaperone BAG1. In line, overexpression of BAG1 increases the proteasome-mediated clearance of these misfolded proteins. Our data suggest that when the misfolded proteins cannot be efficiently transported toward the perinuclear region of the cells, where they are either degraded by autophagy or stored into the aggresome, the cells activate a compensatory mechanism that relies on the induction of BAG1 to target the HSPA8-bound cargo to the proteasome in a dynein-independent manner.

  1. CHOBIMALT: A Cholesterol-Based Detergent†

    Science.gov (United States)

    Howell, Stanley C.; Mittal, Ritesh; Huang, Lijun; Travis, Benjamin; Breyer, Richard M.; Sanders, Charles R.

    2010-01-01

    Cholesterol and its hemisuccinate and sulfate derivatives are widely used in studies of purified membrane proteins, but are difficult to solubilize in aqueous solution, even in the presence of detergent micelles. Other cholesterol derivatives do not form conventional micelles and lead to viscous solutions. To address these problems a cholesterol-based detergent, CHOBIMALT, has been synthesized and characterized. At concentrations above 3–4μM, CHOBIMALT forms micelles without the need for elevated temperatures or sonic disruption. Diffusion and fluorescence measurements indicated that CHOBIMALT micelles are large (210 ± 30 kDa). The ability to solubilize a functional membrane protein was explored using a G-protein coupled receptor, the human kappa opioid receptor type 1 (hKOR1). While CHOBIMALT alone was not found to be effective as a surfactant for membrane extraction, when added to classical detergent micelles CHOBIMALT was observed to dramatically enhance the thermal stability of solubilized hKOR1. PMID:20919740

  2. Bile acid sequestrants for cholesterol

    Science.gov (United States)

    ... ency/patientinstructions/000787.htm Bile acid sequestrants for cholesterol To use the sharing features on this page, ... are medicines that help lower your LDL (bad) cholesterol . Too much cholesterol in your blood can stick ...

  3. CHARACTERIZATION OF A BINDING PROTEIN-DEPENDENT GLUTAMATE TRANSPORT-SYSTEM OF RHODOBACTER-SPHAEROIDES

    NARCIS (Netherlands)

    Jacobs, M.H J; Driessen, A.J.M.; Konings, W.N

    The mechanism of L-glutamate uptake was studied in Rhodobacter sphaeroides. Uptake of L-glutamate is mediated by a high-affinity (K-t of 1.2 mu M), shock-sensitive transport system that is inhibited by vanadate and dependent on the internal pH. From the shock fluid, an L-glutamate-binding protein

  4. Characterization of a Binding Protein-Dependent Glutamate Transport System of Rhodobacter sphaeroides

    NARCIS (Netherlands)

    Jacobs, Mariken H.J.; Driessen, Arnold J.M.; Konings, Wil N.

    1995-01-01

    The mechanism of L-glutamate uptake was studied in Rhodobacter sphaeroides. Uptake of L-glutamate is mediated by a high-affinity (Kt of 1.2 µM), shock-sensitive transport system that is inhibited by vanadate and dependent on the internal pH. From the shock fluid, an L-glutamate-binding protein was

  5. Characterization of drug transport by the human multidrug resistance protein 3 (ABCC3)

    NARCIS (Netherlands)

    Zelcer, N.; Saeki, T.; Reid, G.; Beijnen, J. H.; Borst, P.

    2001-01-01

    We have characterized the substrate specificity and mechanism of transport of the human multidrug resistance-associated protein 3 (MRP3). A murine fibroblast-like cell line generated from the kidneys of mice that lack Mdr1a/b and Mrp1 was retrovirally transduced with MRP3 cDNA. Stable clones

  6. Acid-base status determines the renal expression of Ca2+ and Mg2+ transport proteins.

    NARCIS (Netherlands)

    Nijenhuis, T.; Renkema, K.Y.R.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2006-01-01

    Chronic metabolic acidosis results in renal Ca2+ and Mg2+ wasting, whereas chronic metabolic alkalosis is known to exert the reverse effects. It was hypothesized that these adaptations are mediated at least in part by the renal Ca2+ and Mg2+ transport proteins. The aim of this study, therefore, was

  7. Lipid Raft-Based Membrane Compartmentation of a Plant Transport Protein Expressed in Saccharomyces cerevisiae

    Czech Academy of Sciences Publication Activity Database

    Grossmann, Q.; Opekarová, Miroslava; Nováková, L.; Stolz, J.; Tanner, W.

    2006-01-01

    Roč. 5, č. 6 (2006), s. 945-953 ISSN 1535-9778 R&D Projects: GA MŠk LC545 Institutional research plan: CEZ:AV0Z50200510 Keywords : saccharomyces cerevisiae * plant transport protein * hup1 Subject RIV: EE - Microbiology, Virology Impact factor: 3.707, year: 2006

  8. Disparate effects of p24alpha and p24delta on secretory protein transport and processing.

    Directory of Open Access Journals (Sweden)

    Jeroen R P M Strating

    Full Text Available BACKGROUND: The p24 family is thought to be somehow involved in endoplasmic reticulum (ER-to-Golgi protein transport. A subset of the p24 proteins (p24alpha(3, -beta(1, -gamma(3 and -delta(2 is upregulated when Xenopus laevis intermediate pituitary melanotrope cells are physiologically activated to produce vast amounts of their major secretory cargo, the prohormone proopiomelanocortin (POMC. METHODOLOGY/PRINCIPAL FINDINGS: Here we find that transgene expression of p24alpha(3 or p24delta(2 specifically in the Xenopus melanotrope cells in both cases causes an effective displacement of the endogenous p24 proteins, resulting in severely distorted p24 systems and disparate melanotrope cell phenotypes. Transgene expression of p24alpha(3 greatly reduces POMC transport and leads to accumulation of the prohormone in large, ER-localized electron-dense structures, whereas p24delta(2-transgenesis does not influence the overall ultrastructure of the cells nor POMC transport and cleavage, but affects the Golgi-based processes of POMC glycomaturation and sulfation. CONCLUSIONS/SIGNIFICANCE: Transgenic expression of two distinct p24 family members has disparate effects on secretory pathway functioning, illustrating the specificity and non-redundancy of our transgenic approach. We conclude that members of the p24 family furnish subcompartments of the secretory pathway with specific sets of machinery cargo to provide the proper microenvironments for efficient and correct secretory protein transport and processing.

  9. Structural and functional studies of conserved nucleotide-binding protein LptB in lipopolysaccharide transport

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhongshan [Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, NR4 7TJ (United Kingdom); College of Life Sciences, Sichuan University, Chengdu 610065 (China); Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); Xiang, Quanju [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); Department of Microbiology, College of Resource and Environment Science, Sichuan Agriculture University, Yaan 625000 (China); Zhu, Xiaofeng [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Dong, Haohao [Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); He, Chuan [School of Electronics and Information, Wuhan Technical College of Communications, No. 6 Huangjiahu West Road, Hongshan District, Wuhan, Hubei 430065 (China); Wang, Haiyan; Zhang, Yizheng [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Wang, Wenjian, E-mail: Wenjian166@gmail.com [Laboratory of Department of Surgery, The First Affiliated Hospital, Sun Yat-sen University, 58 Zhongshan Road II, Guangzhou, Guangdong 510080 (China); Dong, Changjiang, E-mail: C.Dong@uea.ac.uk [Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, NR4 7TJ (United Kingdom)

    2014-09-26

    Highlights: • Determination of the structure of the wild-type LptB in complex with ATP and Mg{sup 2+}. • Demonstrated that ATP binding residues are essential for LptB’s ATPase activity and LPS transport. • Dimerization is required for the LptB’s function and LPS transport. • Revealed relationship between activity of the LptB and the vitality of E. coli cells. - Abstract: Lipopolysaccharide (LPS) is the main component of the outer membrane of Gram-negative bacteria, which plays an essential role in protecting the bacteria from harsh conditions and antibiotics. LPS molecules are transported from the inner membrane to the outer membrane by seven LPS transport proteins. LptB is vital in hydrolyzing ATP to provide energy for LPS transport, however this mechanism is not very clear. Here we report wild-type LptB crystal structure in complex with ATP and Mg{sup 2+}, which reveals that its structure is conserved with other nucleotide-binding proteins (NBD). Structural, functional and electron microscopic studies demonstrated that the ATP binding residues, including K42 and T43, are crucial for LptB’s ATPase activity, LPS transport and the vitality of Escherichia coli cells with the exceptions of H195A and Q85A; the H195A mutation does not lower its ATPase activity but impairs LPS transport, and Q85A does not alter ATPase activity but causes cell death. Our data also suggest that two protomers of LptB have to work together for ATP hydrolysis and LPS transport. These results have significant impacts in understanding the LPS transport mechanism and developing new antibiotics.

  10. Structural and functional studies of conserved nucleotide-binding protein LptB in lipopolysaccharide transport

    International Nuclear Information System (INIS)

    Wang, Zhongshan; Xiang, Quanju; Zhu, Xiaofeng; Dong, Haohao; He, Chuan; Wang, Haiyan; Zhang, Yizheng; Wang, Wenjian; Dong, Changjiang

    2014-01-01

    Highlights: • Determination of the structure of the wild-type LptB in complex with ATP and Mg 2+ . • Demonstrated that ATP binding residues are essential for LptB’s ATPase activity and LPS transport. • Dimerization is required for the LptB’s function and LPS transport. • Revealed relationship between activity of the LptB and the vitality of E. coli cells. - Abstract: Lipopolysaccharide (LPS) is the main component of the outer membrane of Gram-negative bacteria, which plays an essential role in protecting the bacteria from harsh conditions and antibiotics. LPS molecules are transported from the inner membrane to the outer membrane by seven LPS transport proteins. LptB is vital in hydrolyzing ATP to provide energy for LPS transport, however this mechanism is not very clear. Here we report wild-type LptB crystal structure in complex with ATP and Mg 2+ , which reveals that its structure is conserved with other nucleotide-binding proteins (NBD). Structural, functional and electron microscopic studies demonstrated that the ATP binding residues, including K42 and T43, are crucial for LptB’s ATPase activity, LPS transport and the vitality of Escherichia coli cells with the exceptions of H195A and Q85A; the H195A mutation does not lower its ATPase activity but impairs LPS transport, and Q85A does not alter ATPase activity but causes cell death. Our data also suggest that two protomers of LptB have to work together for ATP hydrolysis and LPS transport. These results have significant impacts in understanding the LPS transport mechanism and developing new antibiotics

  11. Arctigenin promotes cholesterol efflux from THP-1 macrophages through PPAR-γ/LXR-α signaling pathway.

    Science.gov (United States)

    Xu, Xiaolin; Li, Qian; Pang, Liewen; Huang, Guoqian; Huang, Jiechun; Shi, Meng; Sun, Xiaotian; Wang, Yiqing

    2013-11-15

    Cholesterol efflux from macrophages is a critical mechanism to prevent the development of atherosclerosis. Here, we sought to investigate the effects of arctigenin, a bioactive component of Arctium lappa, on the cholesterol efflux in oxidized low-density lipoprotein (oxLDL)-loaded THP-1 macrophages. Our data showed that arctigenin significantly accelerated apolipoprotein A-I- and high-density lipoprotein-induced cholesterol efflux in both dose- and time-dependent manners. Moreover, arctigenin treatment enhanced the expression of ATP binding cassette transporter A1 (ABCA1), ABCG1, and apoE, all of which are key molecules in the initial step of cholesterol efflux, at both mRNA and protein levels. Arctigenin also caused a concentration-dependent elevation in the expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ) and liver X receptor-alpha (LXR-α). The arctigenin-mediated induction of ABCA1, ABCG1, and apoE was abolished by specific inhibition of PPAR-γ or LXR-α using small interfering RNA technology. Our results collectively indicate that arctigenin promotes cholesterol efflux in oxLDL-loaded THP-1 macrophages through upregulation of ABCA1, ABCG1 and apoE, which is dependent on the enhanced expression of PPAR-γ and LXR-α. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Curcumin inhibits cholesterol uptake in Caco-2 cells by down-regulation of NPC1L1 expression

    Directory of Open Access Journals (Sweden)

    Duan Rui-Dong

    2010-04-01

    Full Text Available Abstract Background Curcumin is a polyphenol and the one of the principle curcuminoids of the spice turmeric. Its antioxidant, anti-cancer and anti-inflammatory effects have been intensively studied. Previous in vivo studies showed that administration of curcumin also decreased cholesterol levels in the blood, and the effects were considered to be related to upregulation of LDL receptor. However, since plasma cholesterol levels are also influenced by the uptake of cholesterol in the gut, which is mediated by a specific transporter Niemann-Pick Cl-like 1 (NPC1L1 protein, the present study is to investigate whether curcumin affects cholesterol uptake in the intestinal Caco-2 cells. Methods Caco-2 cells were cultured to confluence. The micelles composed of bile salt, monoolein, and 14C-cholesterol were prepared. We first incubated the cells with the micelles in the presence and absence of ezetimibe, the specific inhibitor of NPC1L1, to see whether the uptake of the cholesterol in the cells was mediated by NPC1L1. We then pretreated the cells with curcumin at different concentrations for 24 h followed by examination of the changes of cholesterol uptake in these curcumin-treated cells. Finally we determined whether curcumin affects the expression of NPC1L1 by both Western blot analysis and qPCR quantification. Results We found that the uptake of radioactive cholesterol in Caco-2 cells was inhibited by ezetimibe in a dose-dependent manner. The results indicate that the uptake of cholesterol in this study was mediated by NPC1L1. We then pretreated the cells with 25-100 μM curcumin for 24 h and found that such a treatment dose-dependently inhibited cholesterol uptake with 40% inhibition obtained by 100 μM curcumin. In addition, we found that the curcumin-induced inhibition of cholesterol uptake was associated with significant decrease in the levels of NPC1L1 protein and NPC1L1 mRNA, as analyzed by Western blot and qPCR, respectively. Conclusion

  13. Curcumin inhibits cholesterol uptake in Caco-2 cells by down-regulation of NPC1L1 expression.

    Science.gov (United States)

    Feng, Dan; Ohlsson, Lena; Duan, Rui-Dong

    2010-04-19

    Curcumin is a polyphenol and the one of the principle curcuminoids of the spice turmeric. Its antioxidant, anti-cancer and anti-inflammatory effects have been intensively studied. Previous in vivo studies showed that administration of curcumin also decreased cholesterol levels in the blood, and the effects were considered to be related to upregulation of LDL receptor. However, since plasma cholesterol levels are also influenced by the uptake of cholesterol in the gut, which is mediated by a specific transporter Niemann-Pick Cl-like 1 (NPC1L1) protein, the present study is to investigate whether curcumin affects cholesterol uptake in the intestinal Caco-2 cells. Caco-2 cells were cultured to confluence. The micelles composed of bile salt, monoolein, and 14C-cholesterol were prepared. We first incubated the cells with the micelles in the presence and absence of ezetimibe, the specific inhibitor of NPC1L1, to see whether the uptake of the cholesterol in the cells was mediated by NPC1L1. We then pretreated the cells with curcumin at different concentrations for 24 h followed by examination of the changes of cholesterol uptake in these curcumin-treated cells. Finally we determined whether curcumin affects the expression of NPC1L1 by both Western blot analysis and qPCR quantification. We found that the uptake of radioactive cholesterol in Caco-2 cells was inhibited by ezetimibe in a dose-dependent manner. The results indicate that the uptake of cholesterol in this study was mediated by NPC1L1. We then pretreated the cells with 25-100 muM curcumin for 24 h and found that such a treatment dose-dependently inhibited cholesterol uptake with 40% inhibition obtained by 100 muM curcumin. In addition, we found that the curcumin-induced inhibition of cholesterol uptake was associated with significant decrease in the levels of NPC1L1 protein and NPC1L1 mRNA, as analyzed by Western blot and qPCR, respectively. Curcumin inhibits cholesterol uptake through suppression of NPC1L1

  14. Relation between hepatic expression of ATP-binding cassette transporters G5 and G8 and biliary cholesterol secretion in mice

    NARCIS (Netherlands)

    Kosters, A; Frijters, RJJM; Schaap, FG; Vink, E; Plosch, T; Ottenhoff, R; Jirsa, M; De Cuyper, IM; Kuipers, F; Groen, AK

    Background/Aims: Mutations in genes encoding the ATP-binding cassette (ABC)-transporters ABCG5 and ABCG8 underlie sitosterolemia, which is characterized by elevated plasma levels of phytosterols due to increased intestinal absorption and impaired biliary secretion of sterols. The aim of our study

  15. Relation between hepatic expression of ATP-binding cassette transporters G5 and G8 and biliary cholesterol secretion in mice

    NARCIS (Netherlands)

    Kosters, Astrid; Frijters, Raoul J. J. M.; Schaap, Frank G.; Vink, Edwin; Plösch, Torsten; Ottenhoff, Roelof; Jirsa, Milan; de Cuyper, Iris M.; Kuipers, Folkert; Groen, Albert K.

    2003-01-01

    Background/Aims: Mutations in genes encoding the ATP-binding cassette (ABC)-transporters ABCG5 and ABCG8 underlie sitosterolemia, which is characterized by elevated plasma levels of phytosterols due to increased intestinal absorption and impaired biliary secretion of sterols. The aim of our study

  16. Taurine ameliorates cholesterol metabolism by stimulating bile acid production in high-cholesterol-fed rats.

    Science.gov (United States)

    Murakami, Shigeru; Fujita, Michiko; Nakamura, Masakazu; Sakono, Masanobu; Nishizono, Shoko; Sato, Masao; Imaizumi, Katsumi; Mori, Mari; Fukuda, Nobuhiro

    2016-03-01

    This study was designed to investigate the effects of dietary taurine on cholesterol metabolism in high-cholesterol-fed rats. Male Sprague-Dawley rats were randomly divided into two dietary groups (n = 6 in each group): a high-cholesterol diet containing 0.5% cholesterol and 0.15% sodium cholate, and a high-cholesterol diet with 5% (w/w) taurine. The experimental diets were given for 2 weeks. Taurine supplementation reduced the serum and hepatic cholesterol levels by 37% and 32%, respectively. Faecal excretion of bile acids was significantly increased in taurine-treated rats, compared with untreated rats. Biliary bile acid concentrations were also increased by taurine. Taurine supplementation increased taurine-conjugated bile acids by 61% and decreased glycine-conjugated bile acids by 53%, resulting in a significant decrease in the glycine/taurine (G/T) ratio. Among the taurine-conjugated bile acids, cholic acid and deoxycholic acid were significantly increased. In the liver, taurine supplementation increased the mRNA expression and enzymatic activity of hepatic cholesterol 7α-hydroxylase (CYP7A1), the rate-limiting enzyme for bile acid synthesis, by three- and two-fold, respectively. Taurine also decreased the enzymatic activity of acyl-CoA:cholesterol acyltransferase (ACAT) and microsomal triglyceride transfer protein (MTP). These observations suggest that taurine supplementation increases the synthesis and excretion of taurine-conjugated bile acids and stimulates the catabolism of cholesterol to bile acid by elevating the expression and activity of CYP7A1. This may reduce cholesterol esterification and lipoprotein assembly for very low density lipoprotein (VLDL) secretion, leading to reductions in the serum and hepatic cholesterol levels. © 2016 John Wiley & Sons Australia, Ltd.

  17. Tubule urate and PAH transport: sensitivity and specificity of serum protein inhibition

    International Nuclear Information System (INIS)

    Grantham, J.J.; Kennedy, J.; Cowley, B.

    1987-01-01

    Macromolecules in rabbit serum inhibit the cellular uptake and transepithelial secretion of [ 14 C]urate and p-[ 3 H]aminohippurate ([ 3 H]PAH) in rabbit S 2 proximal tubule segments. To understand better the potential role these inhibitors may have in the regulation of renal organic anion excretion, the authors examined the specificity and relative inhibitory effects on tubule urate and PAH transport of albumin and γ-globulin, the major inhibitory proteins in rabbit serum. Native rabbit serum markedly inhibited the cellular accumulation or urate and PAH by isolated nonperfused segments. Urate and PAH transport was also inhibited by bovine serum, human serum, Cohn-fractionated rabbit albumin, and rabbit γ-globulin, but not by Cohn-fractionated bovine serum albumin. α-Lactalbumin and β-lactoglobulin, derived from milk, also inhibited urate and PAH transport, but to a lesser extent than albumin and γ-globulin. The transport inhibitory effects of proteins were independent of their binding to urate and PAH. Unidirectional influx and the steady-state intracellular accumulation of urate and PAH in suspensions of proximal tubules were decreased by rabbit serum proteins, suggesting that these inhibitors act on the external face of the cells to diminish the uptake of the organic anions. These studies indicate that the principal plasma proteins (albumin and γ-globulin) significantly inhibit urate and PAH transporters in the basolateral membranes of S 2 proximal tubules. They suggest that circulating plasma proteins that can penetrate the basement membrane of proximal tubules may directly modulate the renal excretion of urate and PAH

  18. Arctigenin promotes cholesterol efflux from THP-1 macrophages through PPAR-γ/LXR-α signaling pathway

    International Nuclear Information System (INIS)

    Xu, Xiaolin; Li, Qian; Pang, Liewen; Huang, Guoqian; Huang, Jiechun; Shi, Meng; Sun, Xiaotian; Wang, Yiqing

    2013-01-01

    Highlights: •Arctigenin enhanced cholesterol efflux in oxLDL-loaded THP-1 macrophages. •The expression of ABCA1, ABCG1 and apoE was upregulated in arctigenin-treated cells. •Arctigenin promoted the expression of PPAR-γ and LXR-α. •Inhibition of PPAR-γ or LXR-α reversed arctigenin-mediated biological effects. •Arctigenin promotes cholesterol efflux via activation of PPAR-γ/LXR-α/ABCA1 pathway. -- Abstract: Cholesterol efflux from macrophages is a critical mechanism to prevent the development of atherosclerosis. Here, we sought to investigate the effects of arctigenin, a bioactive component of Arctium lappa, on the cholesterol efflux in oxidized low-density lipoprotein (oxLDL)-loaded THP-1 macrophages. Our data showed that arctigenin significantly accelerated apolipoprotein A-I- and high-density lipoprotein-induced cholesterol efflux in both dose- and time-dependent manners. Moreover, arctigenin treatment enhanced the expression of ATP binding cassette transporter A1 (ABCA1), ABCG1, and apoE, all of which are key molecules in the initial step of cholesterol efflux, at both mRNA and protein levels. Arctigenin also caused a concentration-dependent elevation in the expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ) and liver X receptor-alpha (LXR-α). The arctigenin-mediated induction of ABCA1, ABCG1, and apoE was abolished by specific inhibition of PPAR-γ or LXR-α using small interfering RNA technology. Our results collectively indicate that arctigenin promotes cholesterol efflux in oxLDL-loaded THP-1 macrophages through upregulation of ABCA1, ABCG1 and apoE, which is dependent on the enhanced expression of PPAR-γ and LXR-α

  19. Arctigenin promotes cholesterol efflux from THP-1 macrophages through PPAR-γ/LXR-α signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Xiaolin [Department of Cardiothoracic Surgery, Huashan Hospital, Fudan University, Shanghai 200032 (China); Li, Qian [Department of Integrative Medicine and Neurobiology, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai (China); Pang, Liewen; Huang, Guoqian; Huang, Jiechun; Shi, Meng; Sun, Xiaotian [Department of Cardiothoracic Surgery, Huashan Hospital, Fudan University, Shanghai 200032 (China); Wang, Yiqing, E-mail: yiqingwangbiopaper@163.com [Department of Cardiothoracic Surgery, Huashan Hospital, Fudan University, Shanghai 200032 (China)

    2013-11-15

    Highlights: •Arctigenin enhanced cholesterol efflux in oxLDL-loaded THP-1 macrophages. •The expression of ABCA1, ABCG1 and apoE was upregulated in arctigenin-treated cells. •Arctigenin promoted the expression of PPAR-γ and LXR-α. •Inhibition of PPAR-γ or LXR-α reversed arctigenin-mediated biological effects. •Arctigenin promotes cholesterol efflux via activation of PPAR-γ/LXR-α/ABCA1 pathway. -- Abstract: Cholesterol efflux from macrophages is a critical mechanism to prevent the development of atherosclerosis. Here, we sought to investigate the effects of arctigenin, a bioactive component of Arctium lappa, on the cholesterol efflux in oxidized low-density lipoprotein (oxLDL)-loaded THP-1 macrophages. Our data showed that arctigenin significantly accelerated apolipoprotein A-I- and high-density lipoprotein-induced cholesterol efflux in both dose- and time-dependent manners. Moreover, arctigenin treatment enhanced the expression of ATP binding cassette transporter A1 (ABCA1), ABCG1, and apoE, all of which are key molecules in the initial step of cholesterol efflux, at both mRNA and protein levels. Arctigenin also caused a concentration-dependent elevation in the expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ) and liver X receptor-alpha (LXR-α). The arctigenin-mediated induction of ABCA1, ABCG1, and apoE was abolished by specific inhibition of PPAR-γ or LXR-α using small interfering RNA technology. Our results collectively indicate that arctigenin promotes cholesterol efflux in oxLDL-loaded THP-1 macrophages through upregulation of ABCA1, ABCG1 and apoE, which is dependent on the enhanced expression of PPAR-γ and LXR-α.

  20. PXR induces CYP27A1 and regulates cholesterol metabolism in the intestine.

    Science.gov (United States)

    Li, Tiangang; Chen, Wenling; Chiang, John Y L

    2007-02-01

    Mitochondrial sterol 27-hydroxylase (CYP27A1) catalyzes oxidative cleavage of the sterol side chain in the bile acid biosynthetic pathway in the liver and 27-hydroxylation of cholesterol in most tissues. Recent studies suggest that 27-hydroxycholesterol (27-HOC) activates liver orphan receptor alpha (LXRalpha) and induces the cholesterol efflux transporters ABCA1 and ABCG1 in macrophages. The steroid- and bile acid-activated pregnane X receptor (PXR) plays critical roles in the detoxification of bile acids, cholesterol metabolites, and xenobiotics. The role of CYP27A1 in the intestine is not known. This study investigated PXR and CYP27A1 regulation of cholesterol metabolism in the human intestinal cell lines Caco2 and Ls174T. A human PXR ligand, rifampicin, induced CYP27A1 mRNA expression in intestine cells but not in liver cells. Rifampicin induced CYP27A1 gene transcription, increased intracellular 27-HOC levels, and induced ABCA1 and ABCG1 mRNA expression only in intestine cells. A functional PXR binding site was identified in the human CYP27A1 gene. Chromatin immunoprecipitation assays revealed that rifampicin induced the PXR recruitment of steroid receptor coactivator 1 to CYP27A1 chromatin. Cholesterol loading markedly increased intracellular 27-HOC levels in intestine cells. Rifampicin, 27-HOC, and a potent LXRalpha agonist, T0901317, induced ABCA1 and ABCG1 protein expression and stimulated cholesterol efflux from intestine cells to apolipoprotein A-I and HDL. This study suggests an intestine-specific PXR/CYP27A1/LXRalpha pathway that regulates intestine cholesterol efflux and HDL assembly.

  1. Endogenous Cholesterol Excretion Is Negatively Associated With Carotid Intima-Media Thickness in Humans.

    Science.gov (United States)

    Lin, Xiaobo; Racette, Susan B; Ma, Lina; Wallendorf, Michael; Dávila-Román, Victor G; Ostlund, Richard E

    2017-12-01

    Epidemiological studies strongly suggest that lipid factors independent of low-density lipoprotein cholesterol contribute significantly to cardiovascular disease risk. Because circulating lipoproteins comprise only a small fraction of total body cholesterol, the mobilization and excretion of cholesterol from plasma and tissue pools may be an important determinant of cardiovascular disease risk. Our hypothesis is that fecal excretion of endogenous cholesterol is protective against atherosclerosis. Cholesterol metabolism and carotid intima-media thickness were quantitated in 86 nondiabetic adults. Plasma cholesterol was labeled by intravenous infusion of cholesterol-d 7 solubilized in a lipid emulsion and dietary cholesterol by cholesterol-d 5 and the nonabsorbable stool marker sitostanol-d 4 . Plasma and stool samples were collected while subjects consumed a cholesterol- and phytosterol-controlled metabolic kitchen diet and were analyzed by mass spectrometry. Carotid intima-media thickness was negatively correlated with fecal excretion of endogenous cholesterol ( r =-0.426; P cholesterol ( r =-0.472; P ≤0.0001), and daily percent excretion of cholesterol from the rapidly mixing cholesterol pool ( r =-0.343; P =0.0012) and was positively correlated with percent cholesterol absorption ( r =+0.279; P =0.0092). In a linear regression model controlling for age, sex, systolic blood pressure, hemoglobin A1c, low-density lipoprotein, high-density lipoprotein cholesterol, and statin drug use, fecal excretion of endogenous cholesterol remained significant ( P =0.0008). Excretion of endogenous cholesterol is strongly, independently, and negatively associated with carotid intima-media thickness. The reverse cholesterol transport pathway comprising the intestine and the rapidly mixing plasma, and tissue cholesterol pool could be an unrecognized determinant of cardiovascular disease risk not reflected in circulating lipoproteins. Further work is needed to relate measures of

  2. Identification of membrane proteins associated with phenylpropanoid tolerance and transport in Escherichia coli BL21.

    Science.gov (United States)

    Zhou, Jingwen; Wang, Kui; Xu, Sha; Wu, Junjun; Liu, Peiran; Du, Guocheng; Li, Jianghua; Chen, Jian

    2015-01-15

    Phenylpropanoids are phytochemicals produced by some plants and possess a wide variety of biological activities. These compounds exist in plants in low amounts. Production of them in genetically engineered microorganisms has many advantages. A majority of functional phenylpropanoids are toxic to microbial hosts. Export of these compounds may relieve the cellular toxicity and increase the yield. However, proteins and mechanisms involved in phenylpropanoids transport and tolerance remain poorly understood. In this study, 16 membrane proteins that were differentially expressed in Escherichia coli in response to three typical phenylpropanoids (resveratrol, naringenin and rutin) were identified using a membrane proteomics approach. These proteins included outer membrane proteins OmpA, OmpF, OmpW, FadL, TolC, LamB, and YaeT, peripheral membrane proteins AtpD, AtpH, YgaU, OppA, MalK, and MalE, and cytoplasmic membrane proteins OppD, PotG, and ManX. Functions of these proteins were determined by using gene overexpression and silencing. The results suggest that OmpA and FadL may play important roles in the transmembrane export of phenylpropanoids in E. coli. LamB, MalE, MalK and ManX may participate in phenylpropanoid uptake. The role of YgaU in enhancing the tolerance to phenylpropanoids remains to be determined. These results may assist the engineering of microorganisms with enhanced phenylpropanoid producing capabilities. Phenylpropanoids are phytochemicals produced by some plants and possess a wide variety of biological activities. Both the tolerance and the transport of phenylpropanoids play important roles in systematic metabolic engineering of microorganisms to produce these phytochemicals. Both specific and non-specific transporters are essential for these functions but remain poorly understood. This research utilized membrane proteomics to identify E. coli BL21 (DE3) membrane proteins that may be involved in phenylpropanoid transport and tolerance. These results

  3. Axonal transport of enzymes and labeled proteins in experimental axonopathy induced by p-bromophenylacetylurea

    International Nuclear Information System (INIS)

    Jakobsen, J.; Brimijoin, S.

    1981-01-01

    Axonal transport was studied by several techniques in the sciatic nerves of adult male Sprague-Dawley rats with neuropathy induced by treatment with p-bromophenylacetylurea (BPAU) in dimethylsulfoxide solution. Control rats were treated with solvent alone. BPAU, 200 mg/kg, induced severe muscle weakness in the hindlimbs, beginning after a latent period of 1 week and progressing to near total paralysis by 2 weeks. Axonal transport of the endogenous transmitter enzymes, acetylcholinesterase, dopamine-β-hydroxylase and choline acetyltransferase, was normal at both 2 and 15 days after administration of BPAU, as judged by the accumulation of enzyme activity above and below a set of double ligatures on the sciatic nerve. The velocity of fast anterograde transport of [ 35 S]methionine labeled protein was also unaffected by BPAU. However, 4 abnormalities of transport were detected in BPAU treated rats. These abnormalities are discussed. (Auth.)

  4. An egg-enriched diet attenuates plasma lipids and mediates cholesterol metabolism of high-cholesterol fed rats.

    Science.gov (United States)

    Yang, Fang; Ma, Meihu; Xu, Jia; Yu, Xiufang; Qiu, Ning

    2012-03-01

    We investigated the influence of an egg-enriched diet on plasma, hepatic and fecal lipid levels and on gene expression levels of transporters, receptors and enzymes involved in cholesterol metabolism. Sprague-Dawley rats fed an egg-enriched diet had lower plasma triglycerides, total cholesterol, low density lipoprotein (LDL)-cholesterol, hepatic triglyceride, and cholesterol concentrations, and greater plasma high-density lipoprotein cholesterol concentration, fecal neutral sterol and bile acid concentrations than those fed a plain cholesterol diet. Chicken egg yolk had no effect on sterol 12α-hydroxylase and sterol 27α-hydroxylase; but upregulated mRNA levels of hepatic LDL-receptor, cholesterol 7α-hydroxylase (CYP7A1) and lecithin cholesterol acyltransferase, and downregulated hepatic hydroxymethylglutaryl-(HMG)-CoA reductase and acyl-CoA:cholesterol acyltransferase (ACAT) after 90 days. Modification of the lipoprotein profile by an egg-enriched diet was mediated by reducing de novo cholesterol synthesis and enhancing the excretion of fecal cholesterol, via upregulation of CYP7A1 and the LDL receptor, and downregulation of HMG-CoA reductase and ACAT.

  5. Tungsten transport protein A (WtpA) in Pyrococcus furiosus: the first member of a new class of tungstate and molybdate transporters.

    Science.gov (United States)

    Bevers, Loes E; Hagedoorn, Peter-Leon; Krijger, Gerard C; Hagen, Wilfred R

    2006-09-01

    A novel tungstate and molybdate binding protein has been discovered from the hyperthermophilic archaeon Pyrococcus furiosus. This tungstate transport protein A (WtpA) is part of a new ABC transporter system selective for tungstate and molybdate. WtpA has very low sequence similarity with the earlier-characterized transport proteins ModA for molybdate and TupA for tungstate. Its structural gene is present in the genome of numerous archaea and some bacteria. The identification of this new tungstate and molybdate binding protein clarifies the mechanism of tungstate and molybdate transport in organisms that lack the known uptake systems associated with the ModA and TupA proteins, like many archaea. The periplasmic protein of this ABC transporter, WtpA (PF0080), was cloned and expressed in Escherichia coli. Using isothermal titration calorimetry, WtpA was observed to bind tungstate (dissociation constant [K(D)] of 17 +/- 7 pM) and molybdate (K(D) of 11 +/- 5 nM) with a stoichiometry of 1.0 mol oxoanion per mole of protein. These low K(D) values indicate that WtpA has a higher affinity for tungstate than do ModA and TupA and an affinity for molybdate similar to that of ModA. A displacement titration of molybdate-saturated WtpA with tungstate showed that the tungstate effectively replaced the molybdate in the binding site of the protein.

  6. High Blood Cholesterol

    Science.gov (United States)

    ... inflammatory diseases such as psoriasis or to prevent rejection after a transplant Steroids such as prednisone that ... LDL cholesterol levels . Read more Levels of one type of blood fat can signal your risk of developing heart ...

  7. Cholesterol and Women's Health

    Science.gov (United States)

    ... The smallest units of a structure in the body; the building blocks for all parts of the body. Cholesterol: A natural substance that serves as a building block for cells and hormones and helps to ...

  8. Cholesterol synthesis in the lactating cow: Induced expression of candidate genes.

    Science.gov (United States)

    Viturro, Enrique; Koenning, Matthias; Kroemer, Angelika; Schlamberger, Gregor; Wiedemann, Steffi; Kaske, Martin; Meyer, Heinrich H D

    2009-05-01

    Despite the extensive knowledge for other species, cholesterol metabolism in ruminants is nowadays still not clear. Huge differences in milk cholesterol concentration are observed between breeds, managing strategies, individuals and moment of the lactating cycle, but the genetic actors working in the process of cholesterol secretion into milk have not been identified. As ruminant diet contains no cholesterol, understanding the mechanisms and regulation of synthesis, transport and secretion into milk is crucial when trying to reduce the amount of this metabolite in dairy products. The present work aims to study the expression of candidate genes for these processes in the liver of Bos taurus during the lactating cycle. Liver biopsies were obtained from 16 adult brown Swiss cows at different time points (2 weeks pre-partum and 0, 2, 4 and 8 weeks post-partum). After RNA extraction and reverse transcription, gene expression of candidate genes was studied using quantitative RT-PCR. Key enzymes of the cholesterol synthesis (3-hydroxy-methyglutaryl-coenzyme-A (HMG-CoA) synthase, HMG-CoA reductase and farnesyldiphosphat-farnesyltransferase (FDFT)) and gene expression feed-back regulators involved in lipid metabolism (sterol regulatory element binding proteins (SREBP1and 2) SREBP-cleavage activating protein (Scap) were selected as candidate genes. HMG-CoA-reductase and FDFT showed a huge expression increase until week 2 post-partum (pmilk and blood cholesterol levels in B. taurus after parturition might be the result of a coordinated induction in the expression of key liver enzymes and their regulating factors.

  9. Quantitative comparison of the efficacy of various compounds in lowering intracellular cholesterol levels in Niemann-Pick type C fibroblasts.

    Directory of Open Access Journals (Sweden)

    Zachary T Wehrmann

    Full Text Available Niemann-Pick Type C disease (NPC is a lethal, autosomal recessive disorder caused by mutations in the NPC1 and NPC2 cholesterol transport proteins. NPC's hallmark symptoms include an accumulation of unesterified cholesterol and other lipids in the late endosomal and lysosomal cellular compartments, causing progressive neurodegeneration and death. Although the age of onset may vary in those affected, NPC most often manifests in juveniles, and is usually fatal before adolescence. In this study, we investigated the effects of various drugs, many of which modify the epigenetic control of NPC1/NPC2 gene expression, in lowering the otherwise harmful elevated intracellular cholesterol levels in NPC cells. Our studies utilized a previously described image analysis technique, which allowed us to make quantitative comparisons of the efficacy of these drugs in lowering cholesterol levels in a common NPC1 mutant model. Of the drugs analyzed, several that have been previously studied (vorinostat, panobinostat, and β-cyclodextrin significantly lowered the relative amount of unesterified cellular cholesterol, consistent with earlier observations. In addition, a novel potential treatment, rapamycin, likewise alleviated the NPC phenotype. We also studied combinations of effective compounds with β-cyclodextrin; the addition of β-cyclodextrin significantly enhanced the cholesterol-lowering activity of vorinostat and panobinostat, but had mixed effects with rapamycin. Collectively, these results may provide a basis for the eventual development of improved NPC therapies.

  10. Molecular interactions between bile salts, phospholipids and cholesterol : relevance to bile formation, cholesterol crystallization and bile salt toxicity

    NARCIS (Netherlands)

    Moschetta, Antonio

    2001-01-01

    Cholesterol is a nonpolar lipid dietary constituent, absorbed from the small intestine, transported in blood and taken up by the liver. In bile, the sterol is solubilized in mixed micelles by bile salts and phospholipids. In case of supersaturation, cholesterol is kept in vesicles with phospholipid

  11. The role of cholesterol in the association of endoplasmic reticulum membranes with mitochondria

    International Nuclear Information System (INIS)

    Fujimoto, Michiko; Hayashi, Teruo; Su, Tsung-Ping

    2012-01-01

    Highlights: ► The endoplasmic reticulum subdomain termed MAM associates with mitochondria. ► The biophysical role of lipids in the MAM–mitochondria association is unknown. ► The in vitro membrane association assay was used to examine the role of lipids. ► Cholesterol was found to negatively regulate the association. -- Abstract: The unique endoplasmic reticulum (ER) subdomain termed the mitochondria-associated ER membrane (MAM) engages the physical connection between the ER and the mitochondrial outer membrane and plays a role in regulating IP 3 receptor-mediated Ca 2+ influx and the phospholipid transport between the two organelles. The MAM contains certain signaling and membrane-tethering proteins but also lipids including cholesterol. The biophysical role of lipids at the MAM, specifically in the physical interaction between the MAM of the ER and mitochondria, remains not totally clarified. Here we employed the in vitro membrane association assay to investigate the role of cholesterol in the association between MAMs and mitochondria. The purified MAMs and mitochondria were mixed in vitro in a test tube and then the physical association of the two subcellular organelles was quantified indirectly by measuring the presence of the MAM-specific protein sigma-1 receptors in the mitochondria fraction. Purified MAMs contained free cholesterol approximately 7 times higher than that in microsomes. We found that depletion of cholesterol in MAMs with methyl-β-cyclodextrin (MβC) significantly increases the association between MAMs and mitochondria, whereas MβC saturated with cholesterol does not change the association. 14 C-Serine pulse-labeling demonstrated that the treatment of living cells with MβC decreases the level of de novo synthesized 14 C-phosphatidylserine (PtSer) and concomitantly increases greatly the synthesis of 14 C-phosphatidylethanolamine (PtEt). Apparently, cholesterol depletion increased the PtSer transport from MAMs to mitochondria. Our

  12. The role of cholesterol in the association of endoplasmic reticulum membranes with mitochondria

    Energy Technology Data Exchange (ETDEWEB)

    Fujimoto, Michiko [Cellular Stress Signaling Unit, Integrative Neuroscience Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services, Baltimore, MD 21224 (United States); Hayashi, Teruo, E-mail: thayashi@mail.nih.gov [Cellular Stress Signaling Unit, Integrative Neuroscience Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services, Baltimore, MD 21224 (United States); Su, Tsung-Ping, E-mail: tsu@intra.nida.nih.gov [Cellular Pathobiology Section, Integrative Neuroscience Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services, Baltimore, MD 21224 (United States)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer The endoplasmic reticulum subdomain termed MAM associates with mitochondria. Black-Right-Pointing-Pointer The biophysical role of lipids in the MAM-mitochondria association is unknown. Black-Right-Pointing-Pointer The in vitro membrane association assay was used to examine the role of lipids. Black-Right-Pointing-Pointer Cholesterol was found to negatively regulate the association. -- Abstract: The unique endoplasmic reticulum (ER) subdomain termed the mitochondria-associated ER membrane (MAM) engages the physical connection between the ER and the mitochondrial outer membrane and plays a role in regulating IP{sub 3} receptor-mediated Ca{sup 2+} influx and the phospholipid transport between the two organelles. The MAM contains certain signaling and membrane-tethering proteins but also lipids including cholesterol. The biophysical role of lipids at the MAM, specifically in the physical interaction between the MAM of the ER and mitochondria, remains not totally clarified. Here we employed the in vitro membrane association assay to investigate the role of cholesterol in the association between MAMs and mitochondria. The purified MAMs and mitochondria were mixed in vitro in a test tube and then the physical association of the two subcellular organelles was quantified indirectly by measuring the presence of the MAM-specific protein sigma-1 receptors in the mitochondria fraction. Purified MAMs contained free cholesterol approximately 7 times higher than that in microsomes. We found that depletion of cholesterol in MAMs with methyl-{beta}-cyclodextrin (M{beta}C) significantly increases the association between MAMs and mitochondria, whereas M{beta}C saturated with cholesterol does not change the association. {sup 14}C-Serine pulse-labeling demonstrated that the treatment of living cells with M{beta}C decreases the level of de novo synthesized {sup 14}C-phosphatidylserine (PtSer) and concomitantly increases greatly the synthesis of

  13. Reference intervals for serum total cholesterol, HDL cholesterol and ...

    African Journals Online (AJOL)

    Reference intervals of total cholesterol, HDL cholesterol and non-HDL cholesterol concentrations were determined on 309 blood donors from an urban and peri-urban population of Botswana. Using non-parametric methods to establish 2.5th and 97.5th percentiles of the distribution, the intervals were: total cholesterol 2.16 ...

  14. Paraquat Resistant1, a Golgi-localized putative transporter protein, is involved in intracellular transport of paraquat.

    Science.gov (United States)

    Li, Jianyong; Mu, Jinye; Bai, Jiaoteng; Fu, Fuyou; Zou, Tingting; An, Fengying; Zhang, Jian; Jing, Hongwei; Wang, Qing; Li, Zhen; Yang, Shuhua; Zuo, Jianru

    2013-05-01

    Paraquat is one of the most widely used herbicides worldwide. In green plants, paraquat targets the chloroplast by transferring electrons from photosystem I to molecular oxygen to generate toxic reactive oxygen species, which efficiently induce membrane damage and cell death. A number of paraquat-resistant biotypes of weeds and Arabidopsis (Arabidopsis thaliana) mutants have been identified. The herbicide resistance in Arabidopsis is partly attributed to a reduced uptake of paraquat through plasma membrane-localized transporters. However, the biochemical mechanism of paraquat resistance remains poorly understood. Here, we report the identification and characterization of an Arabidopsis paraquat resistant1 (par1) mutant that shows strong resistance to the herbicide without detectable developmental abnormalities. PAR1 encodes a putative l-type amino acid transporter protein localized to the Golgi apparatus. Compared with the wild-type plants, the par1 mutant plants show similar efficiency of paraquat uptake, suggesting that PAR1 is not directly responsible for the intercellular uptake of paraquat. However, the par1 mutation caused a reduction in the accumulation of paraquat in the chloroplast, suggesting that PAR1 is involved in the intracellular transport of paraquat into the chloroplast. We identified a PAR1-like gene, OsPAR1, in rice (Oryza sativa). Whereas the overexpression of OsPAR1 resulted in hypersensitivity to paraquat, the knockdown of its expression using RNA interference conferred paraquat resistance on the transgenic rice plants. These findings reveal a unique mechanism by which paraquat is actively transported into the chloroplast and also provide a practical approach for genetic manipulations of paraquat resistance in crops.

  15. A solute-binding protein for iron transport in Streptococcus iniae

    Directory of Open Access Journals (Sweden)

    Li Anxing

    2010-12-01

    Full Text Available Abstract Background Streptococcus iniae (S. iniae is a major pathogen that causes considerable morbidity and mortality in cultured fish worldwide. The pathogen's ability to adapt to the host affects the extent of infection, hence understanding the mechanisms by which S. iniae overcomes physiological stresses during infection will help to identify potential virulence determinants of streptococcal infection. Grow S. iniae under iron-restricted conditions is one approach for identifying host-specific protein expression. Iron plays an important role in many biological processes but it has low solubility under physiological condition. Many microorganisms have been shown to be able to circumvent this nutritional limitation by forming direct contacts with iron-containing proteins through ATP-binding cassette (ABC transporters. The ABC transporter superfamilies constitute many different systems that are widespread among living organisms with different functions, such as ligands translocation, mRNA translation, and DNA repair. Results An ABC transporter system, named as mtsABC (metal transport system was cloned from S. iniae HD-1, and was found to be involved in heme utilization. mtsABC is cotranscribed by three downstream genes, i.e., mtsA, mtsB, and mtsC. In this study, we cloned the first gene of the mtsABC transporter system (mtsA, and purified the corresponding recombinant protein MtsA. The analysis indicated that MtsA is a putative lipoprotein which binds to heme that can serve as an iron source for the microorganism, and is expressed in vivo during Kunming mice infection by S. iniae HD-1. Conclusions This is believed to be the first report on the cloning the ABC transporter lipoprotein from S. iniae genomic DNA. Together, our data suggested that MtsA is associated with heme, and is expressed in vivo during Kunming mice infection by S. iniae HD-1 which indicated that it can be a potential candidate for S. iniae subunit vaccine.

  16. Solitary BioY Proteins Mediate Biotin Transport into Recombinant Escherichia coli

    Science.gov (United States)

    Finkenwirth, Friedrich; Kirsch, Franziska

    2013-01-01

    Energy-coupling factor (ECF) transporters form a large group of vitamin uptake systems in prokaryotes. They are composed of highly diverse, substrate-specific, transmembrane proteins (S units), a ubiquitous transmembrane protein (T unit), and homo- or hetero-oligomeric ABC ATPases. Biotin transporters represent a special case of ECF-type systems. The majority of the biotin-specific S units (BioY) is known or predicted to interact with T units and ABC ATPases. About one-third of BioY proteins, however, are encoded in organisms lacking any recognizable T unit. This finding raises the question of whether these BioYs function as transporters in a solitary state, a feature ascribed to certain BioYs in the past. To address this question in living cells, an Escherichia coli K-12 derivative deficient in biotin synthesis and devoid of its endogenous high-affinity biotin transporter was constructed as a reference strain. This organism is particularly suited for this purpose because components of ECF transporters do not naturally occur in E. coli K-12. The double mutant was viable in media containing either high levels of biotin or a precursor of the downstream biosynthetic path. Importantly, it was nonviable on trace levels of biotin. Eight solitary bioY genes of proteobacterial origin were individually expressed in the reference strain. Each of the BioYs conferred biotin uptake activity on the recombinants, which was inferred from uptake assays with [3H]biotin and growth of the cells on trace levels of biotin. The results underscore that solitary BioY transports biotin across the cytoplasmic membrane. PMID:23836870

  17. Absorption of Vitamin A and Carotenoids by the Enterocyte: Focus on Transport Proteins

    Directory of Open Access Journals (Sweden)

    Emmanuelle Reboul

    2013-09-01

    Full Text Available Vitamin A deficiency is a public health problem in most developing countries, especially in children and pregnant women. It is thus a priority in health policy to improve preformed vitamin A and/or provitamin A carotenoid status in these individuals. A more accurate understanding of the molecular mechanisms of intestinal vitamin A absorption is a key step in this direction. It was long thought that β-carotene (the main provitamin A carotenoid in human diet, and thus all carotenoids, were absorbed by a passive diffusion process, and that preformed vitamin A (retinol absorption occurred via an unidentified energy-dependent transporter. The discovery of proteins able to facilitate carotenoid uptake and secretion by the enterocyte during the past decade has challenged established assumptions, and the elucidation of the mechanisms of retinol intestinal absorption is in progress. After an overview of vitamin A and carotenoid fate during gastro-duodenal digestion, our focus will be directed to the putative or identified proteins participating in the intestinal membrane and cellular transport of vitamin A and carotenoids across the enterocyte (i.e., Scavenger Receptors or Cellular Retinol Binding Proteins, among others. Further progress in the identification of the proteins involved in intestinal transport of vitamin A and carotenoids across the enterocyte is of major importance for optimizing their bioavailability.

  18. ABC proteins protect the human body and maintain optimal health.

    Science.gov (United States)

    Ueda, Kazumitsu

    2011-01-01

    Human MDR1, a multi-drug transporter gene, was isolated as the first of the eukaryote ATP Binding Cassette (ABC) proteins from a multidrug-resistant carcinoma cell line in 1986. To date, over 25 years, many ABC proteins have been found to play important physiological roles by transporting hydrophobic compounds. Defects in their functions cause various diseases, indicating that endogenous hydrophobic compounds, as well as water-soluble compounds, are properly transported by transmembrane proteins. MDR1 transports a large number of structurally unrelated drugs and is involved in their pharmacokinetics, and thus is a key factor in drug interaction. ABCA1, an ABC protein, eliminates excess cholesterol in peripheral cells by generating HDL. Because ABCA1 is a key molecule in cholesterol homeostasis, its function and expression are highly regulated. Eukaryote ABC proteins function on the body surface facing the outside and in organ pathways to adapt to the extracellular environment and protect the body to maintain optimal health.

  19. The Small Protein SgrT Controls Transport Activity of the Glucose-Specific Phosphotransferase System.

    Science.gov (United States)

    Lloyd, Chelsea R; Park, Seongjin; Fei, Jingyi; Vanderpool, Carin K

    2017-06-01

    The bacterial small RNA (sRNA) SgrS has been a fruitful model for discovery of novel RNA-based regulatory mechanisms and new facets of bacterial physiology and metabolism. SgrS is one of only a few characterized dual-function sRNAs. SgrS can control gene expression posttranscriptionally via sRNA-mRNA base-pairing interactions. Its second function is coding for the small protein SgrT. Previous work demonstrated that both functions contribute to relief of growth inhibition caused by glucose-phosphate stress, a condition characterized by disrupted glycolytic flux and accumulation of sugar phosphates. The base-pairing activity of SgrS has been the subject of numerous studies, but the activity of SgrT is less well characterized. Here, we provide evidence that SgrT acts to specifically inhibit the transport activity of the major glucose permease PtsG. Superresolution microscopy demonstrated that SgrT localizes to the cell membrane in a PtsG-dependent manner. Mutational analysis determined that residues in the N-terminal domain of PtsG are important for conferring sensitivity to SgrT-mediated inhibition of transport activity. Growth assays support a model in which SgrT-mediated inhibition of PtsG transport activity reduces accumulation of nonmetabolizable sugar phosphates and promotes utilization of alternative carbon sources by modulating carbon catabolite repression. The results of this study expand our understanding of a basic and well-studied biological problem, namely, how cells coordinate carbohydrate transport and metabolism. Further, this work highlights the complex activities that can be carried out by sRNAs and small proteins in bacteria. IMPORTANCE Sequencing, annotation and investigation of hundreds of bacterial genomes have identified vast numbers of small RNAs and small proteins, the majority of which have no known function. In this study, we explore the function of a small protein that acts in tandem with a well-characterized small RNA during metabolic

  20. Inhibition of epithelial Na+ transport by atriopeptin, protein kinase c, and pertussis toxin

    International Nuclear Information System (INIS)

    Mohrmann, M.; Cantiello, H.F.; Ausiello, D.A.

    1987-01-01

    The authors have recently shown the selective inhibition of an amiloride-sensitive, conductive pathway for Na + by atrial natriuretic peptide and 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) in the renal epithelial cell line, LLC-PK i . Using 22 Na + fluxes, they further investigated the modulation of Na + transport by atrial natriuretic peptide and by agents that increase cGMP production, activate protein kinase c, or modulate guanine nucleotide regulatory protein function. Sodium nitroprusside increases intracellular cGMP concentrations without affecting cAMP concentrations and completely inhibits amiloride-sensitive Na + uptake in a time- and concentration-dependent manner. Oleoyl 2-acetylglycerol and phorbol 12-myristate 13-acetate, activators of protein kinase c, inhibit Na + uptake by 93 ± 13 and 51 ± 10%, respectively. Prolonged incubation with phorbol ester results in the downregulation of protein kinase c activity and reduces the inhibitory effect of atrial natriuretic peptide, suggesting that the action of this peptide involves stimulation of protein kinase c. Pertussis toxin, which induces the ADP-ribosylation of a 41-kDa guanine nucleotide regulatory protein in LLC-PK i cells, inhibits 22 Na + influx to the same extent as amiloride. Thus, increasing cGMP, activating protein kinase c, and ADP-ribosylating a guanine nucleotide regulatory protein all inhibit Na + uptake. These events may be sequentially involved in the action of atrial natriuretic peptide

  1. Beyond Competitive Inhibition: Regulation of ABC Transporters by Kinases and Protein-Protein Interactions as Potential Mechanisms of Drug-Drug Interactions.

    Science.gov (United States)

    Crawford, Rebecca R; Potukuchi, Praveen K; Schuetz, Erin G; Schuetz, John D

    2018-03-07

    ATP-binding cassette (ABC) transporters are transmembrane efflux transporters mediating the extrusion of an array of substrates ranging from amino acids and lipids to xenobiotics, and many therapeutic compounds, including anticancer drugs. The ABC transporters are also recognized as important contributors to pharmacokinetics, especially in drug-drug interactions and adverse drug effects. Drugs and xenobiotics, as well as pathological conditions, can influence the transcription of ABC transporters, or modify their activity or intracellular localization. Kinases can affect the aforementioned processes for ABC transporters as do protein interactions. In this review, we focus on the ABC transporters ABCB1, ABCB11, ABCC1, ABCC4 and ABCG2 and illustrate how kinases and protein-protein interactions affect these transporters. The clinical relevance of these factors is currently unknown, however these examples suggest that our understanding of drug-drug interactions will benefit from further knowledge of how kinases and protein-protein interactions affect ABC transporters. The American Society for Pharmacology and Experimental Therapeutics.

  2. HDL (Good), LDL (Bad) Cholesterol and Triglycerides

    Science.gov (United States)

    ... be measured by a blood test. LDL (Bad) Cholesterol LDL cholesterol is called “bad” cholesterol. Think of it ... A high triglyceride level combined with low HDL cholesterol or high LDL cholesterol is linked with fatty buildups in artery ...

  3. A Molecular Mechanism for Copper Transportation to Tyrosinase That Is Assisted by a Metallochaperone, Caddie Protein*

    Science.gov (United States)

    Matoba, Yasuyuki; Bando, Naohiko; Oda, Kosuke; Noda, Masafumi; Higashikawa, Fumiko; Kumagai, Takanori; Sugiyama, Masanori

    2011-01-01

    The Cu(II)-soaked crystal structure of tyrosinase that is present in a complex with a protein, designated “caddie,” which we previously determined, possesses two copper ions at its catalytic center. We had identified two copper-binding sites in the caddie protein and speculated that copper bound to caddie may be transported to the tyrosinase catalytic center. In our present study, at a 1.16–1.58 Å resolution, we determined the crystal structures of tyrosinase complexed with caddie prepared by altering the soaking time of the copper ion and the structures of tyrosinase complexed with different caddie mutants that display little or no capacity to activate tyrosinase. Based on these structures, we propose a molecular mechanism by which two copper ions are transported to the tyrosinase catalytic center with the assistance of caddie acting as a metallochaperone. PMID:21730070

  4. Pathobiology of cholesterol gallstone disease: from equilibrium ternary phase diagram to agents preventing cholesterol crystallization and stone formation.

    Science.gov (United States)

    Portincasa, Piero; Moschetta, Antonio; Calamita, Giuseppe; Margari, Antonio; Palasciano, Giuseppe

    2003-03-01

    The primum movens in cholesterol gallstone formation is hypersecretion of hepatic cholesterol, chronic surpersaturation of bile with cholesterol and rapid precipitation of cholesterol crystals in the gallbladder from cholesterol-enriched vesicles. Associated events include biochemical defects (increased biliary mucin, and increased proportions of hydrophobic bile salts in the intestine and gallbladder), motility defects (gallbladder smooth muscle hypocontractility in vitro and gallbladder stasis in vivo, sluggish intestinal transit), and an abnormal genetic background. The study of physical-chemical factors and pathways leading to cholesterol crystallization in bile has clinical relevance and the task can be carried out in different ways. The lithogenicity of bile is investigated in artificial model biles made by three biliary lipids - cholesterol, bile salts and phospholipids - variably combined in systems plotting within the equilibrium ternary phase diagram; also, crystallization propensity of ex vivo incubated human bile is studied by biochemical analysis of precipitated crystals, polarizing quantitative light microscopy and turbidimetric methods. The present review will focus on the recent advances in the field of pathobiology of cholesterol gallstones, by underscoring the role of early events like water transport, lipid transport, crystallization phenomena - including a genetic background - in gallstone pathogenesis. Agents delaying or preventing precipitation of cholesterol crystals and gallstone formation in bile will also be discussed.

  5. Improvement of paracellular transport in the Caco-2 drug screening model using protein-engineered substrates.

    Science.gov (United States)

    DiMarco, Rebecca L; Hunt, Daniel R; Dewi, Ruby E; Heilshorn, Sarah C

    2017-06-01

    The Caco-2 assay has achieved wide popularity among pharmaceutical companies in the past two decades as an in vitro method for estimation of in vivo oral bioavailability of pharmaceutical compounds during preclinical characterization. Despite its popularity, this assay suffers from a severe underprediction of the transport of drugs which are absorbed paracellularly, that is, which pass through the cell-cell tight junctions of the absorptive cells of the small intestine. Here, we propose that simply replacing the collagen I matrix employed in the standard Caco-2 assay with an engineered matrix, we can control cell morphology and hence regulate the cell-cell junctions that dictate paracellular transport. Specifically, we use a biomimetic engineered extracellular matrix (eECM) that contains modular protein domains derived from two ECM proteins found in the small intestine, fibronectin and elastin. This eECM allows us to independently tune the density of cell-adhesive RGD ligands presented to Caco-2 cells as well as the mechanical stiffness of the eECM. We observe that lower amounts of RGD ligand presentation as well as decreased matrix stiffness results in Caco-2 morphologies that more closely resemble primary small intestinal epithelial cells than Caco-2 cells cultured on collagen. Additionally, these matrices result in Caco-2 monolayers with decreased recruitment of actin to the apical junctional complex and increased expression of claudin-2, a tight junction protein associated with higher paracellular permeability that is highly expressed throughout the small intestine. Consistent with these morphological differences, drugs known to be paracellularly transported in vivo exhibited significantly improved transport rates in this modified Caco-2 model. As expected, permeability of transcellularly transported drugs remained unaffected. Thus, we have demonstrated a method of improving the physiological accuracy of the Caco-2 assay that could be readily adopted by

  6. Cytoskeleton-centric protein transportation by exosomes transforms tumor-favorable macrophages

    Science.gov (United States)

    Cui, Yizhi; Zhou, Yanlong; Yin, Xingfeng; Guo, Jiahui; Zhang, Gong; Wang, Tong; He, Qing-Yu

    2016-01-01

    The exosome is a key initiator of pre-metastatic niche in numerous cancers, where macrophages serve as primary inducers of tumor microenvironment. However, the proteome that can be exosomally transported from cancer cells to macrophages has not been sufficiently characterized so far. Here, we used colorectal cancer (CRC) exosomes to educate tumor-favorable macrophages. With a SILAC-based mass spectrometry strategy, we successfully traced the proteome transported from CRC exosomes to macrophages. Such a proteome primarily focused on promoting cytoskeleton rearrangement, which was biologically validated with multiple cell lines. We reproduced the exosomal transportation of functional vimentin as a proof-of-concept example. In addition, we found that some CRC exosomes could be recognized by macrophages via Fc receptors. Therefore, we revealed the active and necessary role of exosomes secreted from CRC cells to transform cancer-favorable macrophages, with the cytoskeleton-centric proteins serving as the top functional unit. PMID:27602764

  7. Maintenance of asymmetric cellular localization of an auxin transport protein through interaction with the actin cytoskeleton

    Science.gov (United States)

    Muday, G. K.

    2000-01-01

    In shoots, polar auxin transport is basipetal (that is, from the shoot apex toward the base) and is driven by the basal localization of the auxin efflux carrier complex. The focus of this article is to summarize the experiments that have examined how the asymmetric distribution of this protein complex is controlled and the significance of this polar distribution. Experimental evidence suggests that asymmetries in the auxin efflux carrier may be established through localized secretion of Golgi vesicles, whereas an attachment of a subunit of the efflux carrier to the actin cytoskeleton may maintain this localization. In addition, the idea that this localization of the efflux carrier may control both the polarity of auxin movement and more globally regulate developmental polarity is explored. Finally, evidence indicating that the gravity vector controls auxin transport polarity is summarized and possible mechanisms for the environmentally induced changes in auxin transport polarity are discussed.

  8. Transportation Stress and Expression of Heat Shock Protein Affecting Pork Quality

    Directory of Open Access Journals (Sweden)

    Shu Tang, , , and

    2014-01-01

    Full Text Available The relationship between heat shock protein (Hsp expression and meat quality were assessed in pigs. Carcasses from 2 h- and 6 h-transported pigs had higher temperatures and lower pH and water holding capacity values in the longissimus dorsi and gluteus maximus superficialis muscles. Long journeys were associated with increased creatine kinase (CK levels. Higher CK levels are indicative of physical stress, as the enzyme is released from muscle fibers as a result of intense muscular exertion. These physiological and enzymatic changes were correlated with increased Hsp70 and decreased Hsp90 expression levels in both skeletal muscles. Animals whose cells contained high levels of Hsp may have had an advantage due to the protective role conferred by Hsp. Reduced Hsp levels were indicative of a higher meat quality and a good welfare of the transported pigs. The stress response declined over time in response to the same stress, such as a 6-h transport stress.

  9. Cytoskeleton-centric protein transportation by exosomes transforms tumor-favorable macrophages.

    Science.gov (United States)

    Chen, Zhipeng; Yang, Lijuan; Cui, Yizhi; Zhou, Yanlong; Yin, Xingfeng; Guo, Jiahui; Zhang, Gong; Wang, Tong; He, Qing-Yu

    2016-10-11

    The exosome is a key initiator of pre-metastatic niche in numerous cancers, where macrophages serve as primary inducers of tumor microenvironment. However, the proteome that can be exosomally transported from cancer cells to macrophages has not been sufficiently characterized so far. Here, we used colorectal cancer (CRC) exosomes to educate tumor-favorable macrophages. With a SILAC-based mass spectrometry strategy, we successfully traced the proteome transported from CRC exosomes to macrophages. Such a proteome primarily focused on promoting cytoskeleton rearrangement, which was biologically validated with multiple cell lines. We reproduced the exosomal transportation of functional vimentin as a proof-of-concept example. In addition, we found that some CRC exosomes could be recognized by macrophages via Fc receptors. Therefore, we revealed the active and necessary role of exosomes secreted from CRC cells to transform cancer-favorable macrophages, with the cytoskeleton-centric proteins serving as the top functional unit.

  10. Cholesterol trafficking and raft-like membrane domain composition mediate scavenger receptor class B type 1-dependent lipid sensing in intestinal epithelial cells.

    Science.gov (United States)

    Morel, Etienne; Ghezzal, Sara; Lucchi, Géraldine; Truntzer, Caroline; Pais de Barros, Jean-Paul; Simon-Plas, Françoise; Demignot, Sylvie; Mineo, Chieko; Shaul, Philip W; Leturque, Armelle; Rousset, Monique; Carrière, Véronique

    2018-02-01

    Scavenger receptor Class B type 1 (SR-B1) is a lipid transporter and sensor. In intestinal epithelial cells, SR-B1-dependent lipid sensing is associated with SR-B1 recruitment in raft-like/ detergent-resistant membrane domains and interaction of its C-terminal transmembrane domain with plasma membrane cholesterol. To clarify the initiating events occurring during lipid sensing by SR-B1, we analyzed cholesterol trafficking and raft-like domain composition in intestinal epithelial cells expressing wild-type SR-B1 or the mutated form SR-B1-Q445A, defective in membrane cholesterol binding and signal initiation. These features of SR-B1 were found to influence both apical cholesterol efflux and intracellular cholesterol trafficking from plasma membrane to lipid droplets, and the lipid composition of raft-like domains. Lipidomic analysis revealed likely participation of d18:0/16:0 sphingomyelin and 16:0/0:0 lysophosphatidylethanolamine in lipid sensing by SR-B1. Proteomic analysis identified proteins, whose abundance changed in raft-like domains during lipid sensing, and these included molecules linked to lipid raft dynamics and signal transduction. These findings provide new insights into the role of SR-B1 in cellular cholesterol homeostasis and suggest molecular links between SR-B1-dependent lipid sensing and cell cholesterol and lipid droplet dynamics. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Specificity of the second binding protein of the peptide ABC-transporter (Dpp) of Lactococcus lactis IL1403

    NARCIS (Netherlands)

    Sanz, Y; Toldra, F; Renault, P; Poolman, B

    2003-01-01

    The genome sequence of Lactococcus lactis IL1403 revealed the presence of a putative peptide-binding protein-dependent ABC-transporter (Dpp). The genes for two peptide-binding proteins (dppA and dppP) precede the membrane components, which include two transmembrane protein genes (dppB and dppC) and

  12. Inclusion of Almonds in a Cholesterol-Lowering Diet Improves Plasma HDL Subspecies and Cholesterol Efflux to Serum in Normal-Weight Individuals with Elevated LDL Cholesterol.

    Science.gov (United States)

    Berryman, Claire E; Fleming, Jennifer A; Kris-Etherton, Penny M

    2017-08-01

    Background : Almonds may increase circulating HDL cholesterol when substituted for a high-carbohydrate snack in an isocaloric diet, yet little is known about the effects on HDL biology and function. Objective: The objective was to determine whether incorporating 43 g almonds/d in a cholesterol-lowering diet would improve HDL subspecies and function, which were secondary study outcomes. Methods: In a randomized, 2-period, crossover, controlled-feeding study, a diet with 43 g almonds/d (percentage of total energy: 51% carbohydrate, 16% protein, and 32% total and 8% saturated fat) was compared with a similar diet with an isocaloric muffin substitution (58% carbohydrate, 15% protein, and 26% total and 8% saturated fat) in men and women with elevated LDL cholesterol. Plasma HDL subspecies and cholesterol efflux from J774 macrophages to human serum were measured at baseline and after each diet period. Diet effects were examined in all participants ( n = 48) and in normal-weight (body mass index: <25; n = 14) and overweight or obese (≥25; n = 34) participants by using linear mixed models. Results: The almond diet, compared with the control diet, increased α-1 HDL [mean ± SEM: 26.7 ± 1.5 compared with 24.3 ± 1.3 mg apolipoprotein A-I (apoA-I)/dL; P = 0.001]. In normal-weight participants, the almond diet, relative to the control diet, increased α-1 HDL (33.7 ± 3.2 compared with 28.4 ± 2.6 mg apoA-I/dL), the α-1 to pre-β-1 ratio [geometric mean (95% CI): 4.3 (3.3, 5.7) compared with 3.1 (2.4, 4.0)], and non-ATP-binding cassette transporter A1 cholesterol efflux (8.3% ± 0.4% compared with 7.8% ± 0.3%) and decreased pre-β-2 (3.8 ± 0.4 compared with 4.6 ± 0.4 mg apoA-I/dL) and α-3 (23.5 ± 0.9 compared with 26.9 ± 1.1 mg apoA-I/dL) HDL ( P < 0.05). No diet effects were observed in the overweight or obese group. Conclusions: Substituting almonds for a carbohydrate-rich snack within a lower-saturated-fat diet may be a simple strategy to maintain a favorable

  13. Protein transport into the human ER and related diseases, Sec61-channelopathies.

    Science.gov (United States)

    Haßdenteufel, Sarah; Klein, Marie-Christine; Melnyk, Armin; Zimmermann, Richard

    2014-12-01

    Protein transport into the human endoplasmic reticulum (ER) is relevant to the biogenesis of most soluble and membrane proteins of organelles, which are involved in endo- or exo-cytsosis. It involves amino-terminal signal peptides in the precursor polypeptides and various transport components in the cytosol plus the ER, and can occur co- or post-translationally. The two mechanisms merge at the level of the ER membrane, specifically at the level of the heterotrimeric Sec61 complex, which forms a dynamic polypeptide-conducting channel in the ER membrane. Since the mammalian ER is also the main intracellular calcium storage organelle, and the Sec61 complex is calcium permeable, the Sec61 complex is tightly regulated in its equilibrium between the closed and open conformations, or "gated", by ligands, such as signal peptides of the transport substrates and the ER lumenal Hsp70-type molecular chaperone BiP. Furthermore, BiP binding to the incoming polypeptide contributes to the efficiency and unidirectionality of transport. Recent insights into the structure and dynamic equilibrium of the Sec61 complex have various mechanistic as well as medical implications.

  14. UV Irradiation Accelerates Amyloid Precursor Protein (APP) Processing and Disrupts APP Axonal Transport

    Science.gov (United States)

    Almenar-Queralt, Angels; Falzone, Tomas L.; Shen, Zhouxin; Lillo, Concepcion; Killian, Rhiannon L.; Arreola, Angela S.; Niederst, Emily D.; Ng, Kheng S.; Kim, Sonia N.; Briggs, Steven P.; Williams, David S.

    2014-01-01

    Overexpression and/or abnormal cleavage of amyloid precursor protein (APP) are linked to Alzheimer's disease (AD) development and progression. However, the molecular mechanisms regulating cellular levels of APP or its processing, and the physiological and pathological consequences of altered processing are not well understood. Here, using mouse and human cells, we found that neuronal damage induced by UV irradiation leads to specific APP, APLP1, and APLP2 decline by accelerating their secretase-dependent processing. Pharmacological inhibition of endosomal/lysosomal activity partially protects UV-induced APP processing implying contribution of the endosomal and/or lysosomal compartments in this process. We found that a biological consequence of UV-induced γ-secretase processing of APP is impairment of APP axonal transport. To probe the functional consequences of impaired APP axonal transport, we isolated and analyzed presumptive APP-containing axonal transport vesicles from mouse cortical synaptosomes using electron microscopy, biochemical, and mass spectrometry analyses. We identified a population of morphologically heterogeneous organelles that contains APP, the secretase machinery, molecular motors, and previously proposed and new residents of APP vesicles. These possible cargoes are enriched in proteins whose dysfunction could contribute to neuronal malfunction and diseases of the nervous system including AD. Together, these results suggest that damage-induced APP processing might impair APP axonal transport, which could result in failure of synaptic maintenance and neuronal dysfunction. PMID:24573290

  15. A mutation in protein phosphatase 2A regulatory subunit A affects auxin transport in Arabidopsis

    Science.gov (United States)

    Garbers, C.; DeLong, A.; Deruere, J.; Bernasconi, P.; Soll, D.; Evans, M. L. (Principal Investigator)

    1996-01-01

    The phytohormone auxin controls processes such as cell elongation, root hair development and root branching. Tropisms, growth curvatures triggered by gravity, light and touch, are also auxin-mediated responses. Auxin is synthesized in the shoot apex and transported through the stem, but the molecular mechanism of auxin transport is not well understood. Naphthylphthalamic acid (NPA) and other inhibitors of auxin transport block tropic curvature responses and inhibit root and shoot elongation. We have isolated a novel Arabidopsis thaliana mutant designated roots curl in NPA (rcn1). Mutant seedlings exhibit altered responses to NPA in root curling and hypocotyl elongation. Auxin efflux in mutant seedlings displays increased sensitivity to NPA. The rcn1 mutation was transferred-DNA (T-DNA) tagged and sequences flanking the T-DNA insert were cloned. Analysis of the RCN1 cDNA reveals that the T-DNA insertion disrupts a gene for the regulatory A subunit of protein phosphatase 2A (PP2A-A). The RCN1 gene rescues the rcn1 mutant phenotype and also complements the temperature-sensitive phenotype of the Saccharomyces cerevisiae PP2A-A mutation, tpd3-1. These data implicate protein phosphatase 2A in the regulation of auxin transport in Arabidopsis.

  16. Aquaporin-11: A channel protein lacking apparent transport function expressed in brain

    Directory of Open Access Journals (Sweden)

    Tsunenari Takashi

    2006-05-01

    Full Text Available Abstract Background The aquaporins are a family of integral membrane proteins composed of two subfamilies: the orthodox aquaporins, which transport only water, and the aquaglyceroporins, which transport glycerol, urea, or other small solutes. Two recently described aquaporins, numbers 11 and 12, appear to be more distantly related to the other mammalian aquaporins and aquaglyceroporins. Results We report on the characterization of Aquaporin-11 (AQP11. AQP11 RNA and protein is found in multiple rat tissues, including kidney, liver, testes and brain. AQP11 has a unique distribution in brain, appearing in Purkinje cell dendrites, hippocampal neurons of CA1 and CA2, and cerebral cortical neurons. Immunofluorescent staining of Purkinje cells indicates that AQP11 is intracellular. Unlike other aquaporins, Xenopus oocytes expressing AQP11 in the plasma membrane failed to transport water, glycerol, urea, or ions. Conclusion AQP11 is functionally distinct from other proteins of the aquaporin superfamily and could represent a new aquaporin subfamily. Further studies are necessary to elucidate the role of AQP11 in the brain.

  17. Coupling Protein Dynamics with Proton Transport in Human Carbonic Anhydrase II.

    Science.gov (United States)

    Taraphder, Srabani; Maupin, C Mark; Swanson, Jessica M J; Voth, Gregory A

    2016-08-25

    The role of protein dynamics in enzyme catalysis is one of the most highly debated topics in enzymology. The main controversy centers around what may be defined as functionally significant conformational fluctuations and how, if at all, these fluctuations couple to enzyme catalyzed events. To shed light on this debate, the conformational dynamics along the transition path surmounting the highest free energy barrier have been herein investigated for the rate limiting proton transport event in human carbonic anhydrase (HCA) II. Special attention has been placed on whether the motion of an excess proton is correlated with fluctuations in the surrounding protein and solvent matrix, which may be rare on the picosecond and subpicosecond time scales of molecular motions. It is found that several active site residues, which do not directly participate in the proton transport event, have a significant impact on the dynamics of the excess proton. These secondary participants are shown to strongly influence the active site environment, resulting in the creation of water clusters that are conducive to fast, moderately slow, or slow proton transport events. The identification and characterization of these secondary participants illuminates the role of protein dynamics in the catalytic efficiency of HCA II.

  18. Enough is enough: TatA demand during Tat-dependent protein transport.

    Science.gov (United States)

    Hauer, René Steffen; Schlesier, René; Heilmann, Kathleen; Dittmar, Julia; Jakob, Mario; Klösgen, Ralf Bernd

    2013-05-01

    The twin-arginine translocation (Tat(1)) pathway is unique with respect to its property to translocate proteins in a fully folded conformation across ion-tight membranes. In chloroplasts and Gram-negative bacteria, Tat translocase consists of the integral subunits TatB and TatC, which are assumed to constitute the membrane receptor, and TatA, a bitopic membrane protein being responsible in a yet unknown manner for the membrane translocation step. Antibody inhibition of intrinsic thylakoidal TatA activity and recovery of transport by heterologously expressed, purified TatA allowed to exactly quantify the amount of TatA required to catalyse membrane transport of the model Tat substrate 16/23. We can show that TatA concentrations in the 100nM range are sufficient to efficiently catalyse membrane transport of the protein, which corresponds well to the amount of TatA identified in thylakoids. Furthermore, TatA shows cooperativity in its catalytic activity suggesting that Tat translocase operates as an allosteric enzyme complex. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Design and engineering of a man-made diffusive electron-transport protein.

    Science.gov (United States)

    Fry, Bryan A; Solomon, Lee A; Leslie Dutton, P; Moser, Christopher C

    2016-05-01

    Maquettes are man-made cofactor-binding oxidoreductases designed from first principles with minimal reference to natural protein sequences. Here we focus on water-soluble maquettes designed and engineered to perform diffusive electron transport of the kind typically carried out by cytochromes, ferredoxins and flavodoxins and other small proteins in photosynthetic and respiratory energy conversion and oxido-reductive metabolism. Our designs were tested by analysis of electron transfer between heme maquettes and the well-known natural electron transporter, cytochrome c. Electron-transfer kinetics were measured from seconds to milliseconds by stopped-flow, while sub-millisecond resolution was achieved through laser photolysis of the carbon monoxide maquette heme complex. These measurements demonstrate electron transfer from the maquette to cytochrome c, reproducing the timescales and charge complementarity modulation observed in natural systems. The ionic strength dependence of inter-protein electron transfer from 9.7×10(6) M(-1) s(-1) to 1.2×10(9) M(-1) s(-1) follows a simple Debye-Hückel model for attraction between +8 net charged oxidized cytochrome c and -19 net charged heme maquette, with no indication of significant protein dipole moment steering. Successfully recreating essential components of energy conversion and downstream metabolism in man-made proteins holds promise for in vivo clinical intervention and for the production of fuel or other industrial products. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. A novel cholesterol-producing Pichia pastoris strain is an ideal host for functional expression of human Na,K-ATPase α3β1 isoform.

    Science.gov (United States)

    Hirz, Melanie; Richter, Gerald; Leitner, Erich; Wriessnegger, Tamara; Pichler, Harald

    2013-11-01

    The heterologous expression of mammalian membrane proteins in lower eukaryotes is often hampered by aberrant protein localization, structure, and function, leading to enhanced degradation and, thus, low expression levels. Substantial quantities of functional membrane proteins are necessary to elucidate their structure-function relationships. Na,K-ATPases are integral, human membrane proteins that specifically interact with cholesterol and phospholipids, ensuring protei