33 results on '"Huijkman N"'
Search Results
2. Prevention of atherosclerosis and hepatic steatosis by combined, liver-specific deletion of GPR146 and ANGPTL3
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Mathada, U. Tharehalli, primary, Rimbert, A., additional, Zhang, B., additional, Kloosterhuis, N., additional, Huijkman, N., additional, Smit, M., additional, Gerding, A., additional, Van De Sluis, B., additional, and Kuivenhoven, J.A., additional
- Published
- 2023
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3. The endosomal sorting protein VPS35 controls lipid homeostasis through regulating hepatic lysosomal function
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Vos, D.Y., primary, Heida, A., additional, Koster, M., additional, Tissink, J., additional, Kuentzel, K.B., additional, Kloosterhuis, N.J., additional, Smit, M., additional, Huijkman, N., additional, Reggiori, F., additional, Mari, M., additional, Scheja, L., additional, Heeren, J., additional, Kratky, D., additional, Kuivenhoven, J., additional, and Van De Sluis, B., additional
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- 2022
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4. Hepatic SMLR1 ablation fully protects against diet-induced atherosclerosis but causes hepatosteatosis via reduced VLDL secretion
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Van Zwol, W., primary, Rimbert, A., additional, Wolters, J.C., additional, Bloks, V., additional, Kloosterhuis, N.J., additional, Huijkman, N., additional, Smit, M., additional, Heeren, J., additional, Scheja, L., additional, Bournez, C.T., additional, Kuipers, J., additional, Rajan, S., additional, Hussain, M., additional, Zimmerman, P., additional, Van De Sluis, B., additional, and Kuivenhoven, J., additional
- Published
- 2022
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5. Homozygous whole body Cbs knockout in adult mice features minimal pathology during ageing despite severe homocysteinemia
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Nakladal, D., primary, Lambooy, S. P. H., additional, Mišúth, S., additional, Čepcová, D., additional, Joschko, C. P., additional, van Buiten, A., additional, Goris, M., additional, Hoogstra‐Berends, F., additional, Kloosterhuis, N. J., additional, Huijkman, N., additional, van de Sluis, B., additional, Diercks, G. F., additional, Buikema, J. H., additional, Henning, R. H., additional, and Deelman, L. E., additional
- Published
- 2022
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6. A novel gene affecting VLDL assembly/secretion
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Van Zwol, W., primary, Rimbert, A., additional, Wolters, K., additional, Larsen, L., additional, Huijkman, N., additional, Kloosterhuis, N., additional, Van De Sluis, B., additional, Zimmermann, P., additional, and Kuivenhoven, J.A., additional
- Published
- 2021
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7. A cargo-specific role for hepatocyte retriever in lipoprotein receptor recycling
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Vos, D.Y., primary, Smit, M., additional, Huijkman, N., additional, Kloosterhuis, N., additional, Wolters, K., additional, Kuivenhoven, J.A., additional, and Van De Sluis, B., additional
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- 2021
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8. The U2-spliceosome and its interactors regulate the levels and activity of the LDL receptor in humans
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Zanoni, P., primary, Panteloglou, G., additional, Othman, A., additional, Haas, J.T., additional, Meier, R., additional, Rimbert, A., additional, Futema, M., additional, Khalil, Y. Abou, additional, Norrelykke, S.F., additional, Rzepiela, A.J., additional, Stoma, S., additional, Stebler, M., additional, Van Dijk, F., additional, Wijers, M., additional, Wolters, J.C., additional, Dalila, N., additional, Huijkman, N., additional, Smit, M., additional, Gallo, A., additional, Carreau, V., additional, Philippi, A., additional, Rabès, J.-P., additional, Boileau, C., additional, Visentin, M., additional, Vonghia, L., additional, Weyler, J., additional, Francque, S.M., additional, Verrijken, A., additional, Verhaegen, A., additional, Van Gaal, L., additional, Van Der Graaf, A., additional, Van Rosmalen, B.V., additional, Velagapudi, S., additional, Yalcinkaya, M., additional, Keel, M., additional, Radosavljevic, S., additional, Geier, A., additional, Tybjærg-Hansen, A., additional, Varret, M., additional, Rohrer, L., additional, Humphries, S.E., additional, Staels, B., additional, Van De Sluis, B., additional, Kuivenhoven, J.A., additional, and Von Eckardstein, A., additional
- Published
- 2020
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9. Rare GPR146 variants and their impact in the regulation of plasma lipid levels
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Velasquez, N. Loaiza, primary, Rimbert, A., additional, Dalila, N., additional, Oldoni, F., additional, Balder, J.W., additional, Huijkman, N., additional, Tybjærg-Hansen, A., additional, Van De Sluis, B., additional, and Kuivenhoven, J.A., additional
- Published
- 2020
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10. The Parkinson’s Disease Gene Vps35 Regulates Plasma Ldl Cholesterol Levels In A Ccc/Wash-Dependent Manner
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Vos, D., primary, Wijers, M., additional, Smit, M., additional, Huijkman, N., additional, Kloosterhuis, N., additional, Wolters, K., additional, Kuivenhoven, J.A., additional, and van de Sluis, B., additional
- Published
- 2019
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11. A Common Variant In Ccdc93 Decreases Ldl-C And Protects Against Myocardial Infarction By Regulating Endosomal Trafficking Of Ldl-Receptor
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Rimbert, A., primary, Dalila, N., additional, Wolters, J., additional, Huijkman, N., additional, Smit, M., additional, Kloosterhuis, N., additional, Riemsma, M., additional, van der Veen, Y., additional, Singla, A., additional, van Dijk, F., additional, Frikke-Schmidt, R., additional, Burtsein, E., additional, Tybjaerg-Hansen, T.H., additional, van de Sluis, B., additional, and Kuivenhoven, J.A., additional
- Published
- 2019
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12. The COMMD Family Regulates Plasma LDL Levels and Attenuates Atherosclerosis Through Stabilizing the CCC Complex in Endosomal LDLR Trafficking
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Fedoseienko, A., Wijers, M., Wolters, J.C., Dekker, D, Smit, M, Huijkman, N., Kloosterhuis, N., Klug, H., Schepers, A., Dijk, K van, Levels, J.H., Billadeau, D.D., Hofker, M.H., Deursen, J. van, Westerterp, M., Burstein, E., Kuivenhoven, J.A., Sluis, B. van de, Fedoseienko, A., Wijers, M., Wolters, J.C., Dekker, D, Smit, M, Huijkman, N., Kloosterhuis, N., Klug, H., Schepers, A., Dijk, K van, Levels, J.H., Billadeau, D.D., Hofker, M.H., Deursen, J. van, Westerterp, M., Burstein, E., Kuivenhoven, J.A., and Sluis, B. van de
- Abstract
Item does not contain fulltext, RATIONALE: COMMD (copper metabolism MURR1 domain)-containing proteins are a part of the CCC (COMMD-CCDC22 [coiled-coil domain containing 22]-CCDC93 [coiled-coil domain containing 93]) complex facilitating endosomal trafficking of cell surface receptors. Hepatic COMMD1 inactivation decreases CCDC22 and CCDC93 protein levels, impairs the recycling of the LDLR (low-density lipoprotein receptor), and increases plasma low-density lipoprotein cholesterol levels in mice. However, whether any of the other COMMD members function similarly as COMMD1 and whether perturbation in the CCC complex promotes atherogenesis remain unclear. OBJECTIVE: The main aim of this study is to unravel the contribution of evolutionarily conserved COMMD proteins to plasma lipoprotein levels and atherogenesis. METHODS AND RESULTS: Using liver-specific Commd1, Commd6, or Commd9 knockout mice, we investigated the relation between the COMMD proteins in the regulation of plasma cholesterol levels. Combining biochemical and quantitative targeted proteomic approaches, we found that hepatic COMMD1, COMMD6, or COMMD9 deficiency resulted in massive reduction in the protein levels of all 10 COMMDs. This decrease in COMMD protein levels coincided with destabilizing of the core (CCDC22, CCDC93, and chromosome 16 open reading frame 62 [C16orf62]) of the CCC complex, reduced cell surface levels of LDLR and LRP1 (LDLR-related protein 1), followed by increased plasma low-density lipoprotein cholesterol levels. To assess the direct contribution of the CCC core in the regulation of plasma cholesterol levels, Ccdc22 was deleted in mouse livers via CRISPR/Cas9-mediated somatic gene editing. CCDC22 deficiency also destabilized the complete CCC complex and resulted in elevated plasma low-density lipoprotein cholesterol levels. Finally, we found that hepatic disruption of the CCC complex exacerbates dyslipidemia and atherosclerosis in ApoE3*Leiden mice. CONCLUSIONS: Collectively, these findings demonstrate a strong inter
- Published
- 2018
13. NF-kappaB p65 serine 467 phosphorylation sensitizes mice to weight gain and TNFalpha-or diet-induced inflammation
- Author
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Riedlinger, T., Dommerholt, M.B., Wijshake, T., Kruit, J.K., Huijkman, N., Dekker, D, Koster, M., Kloosterhuis, N., Koonen, D.P., Bruin, A. de, Baker, D., Hofker, M.H., Deursen, J. van, Jonker, J.W., Schmitz, M.L., Sluis, B. van de, Riedlinger, T., Dommerholt, M.B., Wijshake, T., Kruit, J.K., Huijkman, N., Dekker, D, Koster, M., Kloosterhuis, N., Koonen, D.P., Bruin, A. de, Baker, D., Hofker, M.H., Deursen, J. van, Jonker, J.W., Schmitz, M.L., and Sluis, B. van de
- Abstract
Item does not contain fulltext, The NF-kappaB family of transcription factors is essential for an effective immune response, but also controls cell metabolism, proliferation and apoptosis. Its broad relevance and the high connectivity to diverse signaling pathways require a tight control of NF-kappaB activity. To investigate the control of NF-kappaB activity by phosphorylation of the NF-kappaB p65 subunit, we generated a knock-in mouse model in which serine 467 (the mouse homolog of human p65 serine 468) was replaced with a non-phosphorylatable alanine (S467A). This substitution caused reduced p65 protein synthesis and diminished TNFalpha-induced expression of a selected group of NF-kappaB-dependent genes. Intriguingly, high-fat fed S467A mice displayed increased locomotor activity and energy expenditure, which coincided with a reduced body weight gain. Although glucose metabolism or insulin sensitivity was not improved, diet-induced liver inflammation was diminished in S467A mice. Altogether, this study demonstrates that phosphorylation of p65 serine 467 augment NF-kappaB activity and exacerbates various deleterious effects of overnutrition in mice.
- Published
- 2017
14. Pharmacological activation of LXR in utero directly influences ABC transporter expression and function in mice but does not affect adult cholesterol metabolism
- Author
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van Straten, E. M. E., primary, Huijkman, N. C. A., additional, Baller, J. F. W., additional, Kuipers, F., additional, and Plösch, T., additional
- Published
- 2008
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15. Phylogenetic relationships of carnitine- and choline acyltransferases
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van der Leij, F. R., primary, Huijkman, N. C. A., additional, Kuipers, J. R. G., additional, and Bartelds, B., additional
- Published
- 2001
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16. Selective Hepatic Cbs Knockout Aggravates Liver Damage, Endothelial Dysfunction and ROS Stress in Mice Fed a Western Diet.
- Author
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Lambooy S, Heida A, Joschko C, Nakladal D, van Buiten A, Kloosterhuis N, Huijkman N, Gerding A, van de Sluis B, Henning R, and Deelman L
- Subjects
- Mice, Animals, Reactive Oxygen Species, Diet, Western adverse effects, Hydrogen Peroxide, Mice, Knockout, Liver, Cystathionine beta-Synthase genetics, Diet, High-Fat adverse effects, Mice, Inbred C57BL, Disease Models, Animal, Non-alcoholic Fatty Liver Disease genetics, Hyperhomocysteinemia
- Abstract
Cystathionine-β-synthase (CBS) is highly expressed in the liver, and deficiencies in Cbs lead to hyperhomocysteinemia (HHCy) and disturbed production of antioxidants such as hydrogen sulfide. We therefore hypothesized that liver-specific Cbs deficient (LiCKO) mice would be particularly susceptible to the development of non-alcoholic fatty liver disease (NAFLD). NAFLD was induced by a high-fat high-cholesterol (HFC) diet; LiCKO and controls were split into eight groups based on genotype (con, LiCKO), diet (normal diet, HFC), and diet duration (12 weeks, 20 weeks). LiCKO mice displayed intermediate to severe HHCy. Plasma H
2 O2 was increased by HFC, and further aggravated in LiCKO. LiCKO mice fed an HFC diet had heavier livers, increased lipid peroxidation, elevated ALAT, aggravated hepatic steatosis, and inflammation. LiCKO mice showed decreased L-carnitine in the liver, but this did not result in impaired fatty acid oxidation. Moreover, HFC-fed LiCKO mice demonstrated vascular and renal endothelial dysfunction. Liver and endothelial damage correlated significantly with systemic ROS status. In conclusion, this study demonstrates an important role for CBS in the liver in the development of NAFLD, which is most probably mediated through impaired defense against oxidative stress., Competing Interests: The authors declare no conflict of interest.- Published
- 2023
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17. Cargo-Specific Role for Retriever Subunit VPS26C in Hepatocyte Lipoprotein Receptor Recycling to Control Postprandial Triglyceride-Rich Lipoproteins.
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Vos DY, Wijers M, Smit M, Huijkman N, Kloosterhuis NJ, Wolters JC, Tissink JJ, Pronk ACM, Kooijman S, Rensen PCN, Kuivenhoven JA, and van de Sluis B
- Subjects
- Animals, Humans, Mice, Hepatocytes metabolism, Lipoproteins metabolism, Mice, Knockout, Receptors, LDL, Triglycerides metabolism, Low Density Lipoprotein Receptor-Related Protein-1 genetics, Proprotein Convertase 9 genetics, Proprotein Convertase 9 metabolism
- Abstract
Background: The copper metabolism MURR1 domains/coiled-coil domain containing 22/coiled-coil domain containing 93 (CCC) complex is required for the transport of low-density lipoprotein receptor (LDLR) and LRP1 (LDLR-related protein 1) from endosomes to the cell surface of hepatocytes. Impaired functioning of hepatocytic CCC causes hypercholesterolemia in mice, dogs, and humans. Retriever, a protein complex consisting of subunits VPS26C, VPS35L, and VPS29, is associated with CCC, but its role in endosomal lipoprotein receptor transport is unclear. We here investigated the contribution of retriever to hepatocytic lipoprotein receptor recycling and plasma lipids regulation., Methods: Using somatic CRISPR/Cas9 gene editing, we generated liver-specific VPS35L or VPS26C-deficient mice. We determined total and surface levels of LDLR and LRP1 and plasma lipids. In addition, we studied the protein levels and composition of CCC and retriever., Results: Hepatocyte VPS35L deficiency reduced VPS26C levels but had minimal impact on CCC composition. VPS35L deletion decreased hepatocytic surface expression of LDLR and LRP1, accompanied by a 21% increase in plasma cholesterol levels. Hepatic VPS26C ablation affected neither levels of VPS35L and CCC subunits, nor plasma lipid concentrations. However, VPS26C deficiency increased hepatic LDLR protein levels by 2-fold, probably compensating for reduced LRP1 functioning, as we showed in VPS26C-deficient hepatoma cells. Upon PCSK9 (proprotein convertase subtilisin/kexin type 9)-mediated LDLR elimination, VPS26C ablation delayed postprandial triglyceride clearance and increased plasma triglyceride levels by 26%., Conclusions: Our study suggests that VPS35L is shared between retriever and CCC to facilitate LDLR and LRP1 transport from endosomes to the cell surface. Conversely, retriever subunit VPS26C selectively transports LRP1, but not LDLR, and thereby may control hepatic uptake of postprandial triglyceride-rich lipoprotein remnants.
- Published
- 2023
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18. Butyrate oxidation attenuates the butyrate-induced improvement of insulin sensitivity in myotubes.
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Rios-Morales M, Vieira-Lara MA, Homan E, Langelaar-Makkinje M, Gerding A, Li Z, Huijkman N, Rensen PCN, Wolters JC, Reijngoud DJ, and Bakker BM
- Subjects
- Butyrates metabolism, Butyrates pharmacology, Coenzyme A, Dietary Fiber metabolism, Dietary Fiber pharmacology, Fatty Acids metabolism, Histone Deacetylases genetics, Histone Deacetylases metabolism, Histones metabolism, Humans, Mitochondrial Proteins metabolism, Muscle Fibers, Skeletal metabolism, Palmitates pharmacology, RNA, Messenger metabolism, Receptor, Insulin metabolism, Diabetes Mellitus, Type 2 metabolism, Insulin Resistance physiology, Insulins metabolism, Insulins pharmacology
- Abstract
Skeletal muscle insulin resistance is a key pathophysiological process that precedes the development of type 2 diabetes. Whereas an overload of long-chain fatty acids can induce muscle insulin resistance, butyrate, a short-chain fatty acid (SCFA) produced from dietary fibre fermentation, prevents it. This preventive role of butyrate has been attributed to histone deacetylase (HDAC)-mediated transcription regulation and activation of mitochondrial fatty-acid oxidation. Here we address the interplay between butyrate and the long-chain fatty acid palmitate and investigate how transcription, signalling and metabolism are integrated to result in the butyrate-induced skeletal muscle metabolism remodelling. Butyrate enhanced insulin sensitivity in palmitate-treated, insulin-resistant C2C12 cells, as shown by elevated insulin receptor 1 (IRS1) and pAKT protein levels and Slc2a4 (GLUT4) mRNA, which led to a higher glycolytic capacity. Long-chain fatty-acid oxidation capacity and other functional respiration parameters were not affected. Butyrate did upregulate mitochondrial proteins involved in its own oxidation, as well as concentrations of butyrylcarnitine and hydroyxybutyrylcarnitine. By knocking down the gene encoding medium-chain 3-ketoacyl-CoA thiolase (MCKAT, Acaa2), butyrate oxidation was inhibited, which amplified the effects of the SCFA on insulin sensitivity and glycolysis. This response was associated with enhanced HDAC inhibition, based on histone 3 acetylation levels. Butyrate enhances insulin sensitivity and induces glycolysis, without the requirement of upregulated long-chain fatty acid oxidation. Butyrate catabolism functions as an escape valve that attenuates HDAC inhibition. Thus, inhibition of butyrate oxidation indirectly prevents insulin resistance and stimulates glycolytic flux in myotubes treated with butyrate, most likely via an HDAC-dependent mechanism., Competing Interests: Declaration of competing interest None., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)
- Published
- 2022
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19. Mice with a deficiency in Peroxisomal Membrane Protein 4 (PXMP4) display mild changes in hepatic lipid metabolism.
- Author
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Blankestijn M, Bloks VW, Struik D, Huijkman N, Kloosterhuis N, Wolters JC, Wanders RJA, Vaz FM, Islinger M, Kuipers F, van de Sluis B, Groen AK, Verkade HJ, and Jonker JW
- Subjects
- Animals, Bile Acids and Salts metabolism, CRISPR-Cas Systems, Diet methods, Female, Fenofibrate administration & dosage, Gene Editing methods, Gene Knockout Techniques methods, Male, Membrane Proteins genetics, Mice, Mice, Inbred C57BL, Mice, Knockout, Oxidation-Reduction drug effects, PPAR alpha metabolism, Peroxisomes drug effects, Peroxisomes metabolism, Phytanic Acid metabolism, Phytol administration & dosage, Fatty Acids metabolism, Fatty Acids, Unsaturated metabolism, Liver metabolism, Membrane Proteins metabolism, Signal Transduction genetics
- Abstract
Peroxisomes play an important role in the metabolism of a variety of biomolecules, including lipids and bile acids. Peroxisomal Membrane Protein 4 (PXMP4) is a ubiquitously expressed peroxisomal membrane protein that is transcriptionally regulated by peroxisome proliferator-activated receptor α (PPARα), but its function is still unknown. To investigate the physiological function of PXMP4, we generated a Pxmp4 knockout (Pxmp4
-/- ) mouse model using CRISPR/Cas9-mediated gene editing. Peroxisome function was studied under standard chow-fed conditions and after stimulation of peroxisomal activity using the PPARα ligand fenofibrate or by using phytol, a metabolite of chlorophyll that undergoes peroxisomal oxidation. Pxmp4-/- mice were viable, fertile, and displayed no changes in peroxisome numbers or morphology under standard conditions. Also, no differences were observed in the plasma levels of products from major peroxisomal pathways, including very long-chain fatty acids (VLCFAs), bile acids (BAs), and BA intermediates di- and trihydroxycholestanoic acid. Although elevated levels of the phytol metabolites phytanic and pristanic acid in Pxmp4-/- mice pointed towards an impairment in peroxisomal α-oxidation capacity, treatment of Pxmp4-/- mice with a phytol-enriched diet did not further increase phytanic/pristanic acid levels. Finally, lipidomic analysis revealed that loss of Pxmp4 decreased hepatic levels of the alkyldiacylglycerol class of neutral ether lipids, particularly those containing polyunsaturated fatty acids. Together, our data show that while PXMP4 is not critical for overall peroxisome function under the conditions tested, it may have a role in the metabolism of (ether)lipids., (© 2022. The Author(s).)- Published
- 2022
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20. Corrigendum to "NF-κB p65 serine 467 phosphorylation sensitizes mice to weight gain and TNFα-or diet-induced inflammation" [Biochim. Biophys. Acta Mol. Cell Res., 1864 (2017): 1785-1798].
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Riedlinger T, Dommerholt MB, Wijshake T, Kruit JK, Huijkman N, Dekker D, Koster M, Kloosterhuis N, Koonen DPY, de Bruin A, Baker D, Hofker MH, van Deursen J, Jonker JW, Lienhard Schmitz M, and van de Sluis B
- Published
- 2021
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21. Taking One Step Back in Familial Hypercholesterolemia: STAP1 Does Not Alter Plasma LDL (Low-Density Lipoprotein) Cholesterol in Mice and Humans.
- Author
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Loaiza N, Hartgers ML, Reeskamp LF, Balder JW, Rimbert A, Bazioti V, Wolters JC, Winkelmeijer M, Jansen HPG, Dallinga-Thie GM, Volta A, Huijkman N, Smit M, Kloosterhuis N, Koster M, Svendsen AF, van de Sluis B, Hovingh GK, Grefhorst A, and Kuivenhoven JA
- Subjects
- Animals, Atherosclerosis blood, Atherosclerosis genetics, B-Lymphocytes immunology, Cell Line, Tumor, Disease Models, Animal, Female, Hep G2 Cells, Humans, Lipids blood, Lymphocytes immunology, Male, Mice, Knockout, Monocytes immunology, Adaptor Proteins, Signal Transducing physiology, Cholesterol, LDL blood, Hyperlipoproteinemia Type II blood, Hyperlipoproteinemia Type II genetics
- Abstract
Objective: STAP1 , encoding for STAP1 (signal transducing adaptor family member 1), has been reported as a candidate gene associated with familial hypercholesterolemia. Unlike established familial hypercholesterolemia genes, expression of STAP1 is absent in liver but mainly observed in immune cells. In this study, we set out to validate STAP1 as a familial hypercholesterolemia gene. Approach and Results: A whole-body Stap1 knockout mouse model ( Stap1
-/- ) was generated and characterized, without showing changes in plasma lipid levels compared with controls. In follow-up studies, bone marrow from Stap1-/- mice was transplanted to Ldlr-/- mice, which did not show significant changes in plasma lipid levels or atherosclerotic lesions. To functionally assess whether STAP1 expression in B cells can affect hepatic function, HepG2 cells were cocultured with peripheral blood mononuclear cells isolated from heterozygotes carriers of STAP1 variants and controls. The peripheral blood mononuclear cells from STAP1 variant carriers and controls showed similar LDLR mRNA and protein levels. Also, LDL (low-density lipoprotein) uptake by HepG2 cells did not differ upon coculturing with peripheral blood mononuclear cells isolated from either STAP1 variant carriers or controls. In addition, plasma lipid profiles of 39 carriers and 71 family controls showed no differences in plasma LDL cholesterol, HDL (high-density lipoprotein) cholesterol, triglycerides, and lipoprotein(a) levels. Similarly, B-cell populations did not differ in a group of 10 STAP1 variant carriers and 10 age- and sex-matched controls. Furthermore, recent data from the UK Biobank do not show association between STAP1 rare gene variants and LDL cholesterol., Conclusions: Our combined studies in mouse models and carriers of STAP1 variants indicate that STAP1 is not a familial hypercholesterolemia gene.- Published
- 2020
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22. A human-like bile acid pool induced by deletion of hepatic Cyp2c70 modulates effects of FXR activation in mice.
- Author
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de Boer JF, Verkade E, Mulder NL, de Vries HD, Huijkman N, Koehorst M, Boer T, Wolters JC, Bloks VW, van de Sluis B, and Kuipers F
- Subjects
- Animals, Mice, Mice, Inbred C57BL, Mice, Transgenic, Bile Acids and Salts metabolism, Cytochrome P-450 Enzyme System metabolism, Liver metabolism, Receptors, Cytoplasmic and Nuclear metabolism
- Abstract
Bile acids (BAs) facilitate intestinal absorption of lipid-soluble nutrients and modulate various metabolic pathways through the farnesoid X receptor (FXR) and Takeda G-protein-coupled receptor 5. These receptors are targets for therapy in cholestatic and metabolic diseases. However, dissimilarities in BA metabolism between humans and mice complicate translation of preclinical data. Cytochrome P450 family 2 subfamily c polypeptide 70 (CYP2C70) was recently proposed to catalyze the formation of rodent-specific muricholic acids (MCAs). With CRISPR/Cas9-mediated somatic genome editing, we generated an acute hepatic Cyp2c70 knockout mouse model ( Cyp2c70
ako ) to clarify the role of CYP2C70 in BA metabolism in vivo and evaluate whether its activity modulates effects of pharmacologic FXR activation on cholesterol homeostasis. In Cyp2c70ako mice, chenodeoxycholic acid (CDCA) increased at the expense of βMCA, resulting in a more hydrophobic human-like BA pool. Tracer studies demonstrated that, in vivo, CYP2C70 catalyzes the formation of βMCA primarily by sequential 6β-hydroxylation and C7-epimerization of CDCA, generating αMCA as an intermediate metabolite. Physiologically, the humanized BA composition in Cyp2c70ako mice blunted the stimulation of fecal cholesterol disposal in response to FXR activation compared with WT mice, predominantly due to reduced stimulation of transintestinal cholesterol excretion. Thus, deletion of hepatic Cyp2c70 in adult mice translates into a human-like BA pool composition and impacts the response to pharmacologic FXR activation. This Cyp2c70ako mouse model may be a useful tool for future studies of BA signaling and metabolism that informs human disease development and treatment., (Copyright © 2020 de Boer et al.)- Published
- 2020
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23. A common variant in CCDC93 protects against myocardial infarction and cardiovascular mortality by regulating endosomal trafficking of low-density lipoprotein receptor.
- Author
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Rimbert A, Dalila N, Wolters JC, Huijkman N, Smit M, Kloosterhuis N, Riemsma M, van der Veen Y, Singla A, van Dijk F, Frikke-Schmidt R, Burstein E, Tybjærg-Hansen A, van de Sluis B, and Kuivenhoven JA
- Subjects
- Animals, Cholesterol, LDL genetics, Genome-Wide Association Study, Humans, Mice, Receptors, LDL genetics, Coronary Artery Disease, Myocardial Infarction genetics, Myocardial Infarction prevention & control, Vesicular Transport Proteins genetics
- Abstract
Aims: Genome-wide association studies have previously identified INSIG2 as a candidate gene for plasma low-density lipoprotein cholesterol (LDL-c). However, we suspect a role for CCDC93 in the same locus because of its involvement in the recycling of the LDL-receptor (LDLR)., Methods and Results: Characterization of the INSIG2 locus was followed by studies in over 107 000 individuals from the general population, the Copenhagen General Population Study and the Copenhagen City Heart Study, for associations of genetic variants with plasma lipids levels, with risk of myocardial infarction (MI) and with cardiovascular mortality. CCDC93 was furthermore studied in cells and mice. The lead variant of the INSIG2 locus (rs10490626) is not associated with changes in the expression of nearby genes but is a part of a genetic block, which excludes INSIG2. This block includes a coding variant in CCDC93 p.Pro228Leu, which is in strong linkage disequilibrium with rs10490626 (r2 > 0.96). In the general population, separately and combined, CCDC93 p.Pro228Leu is dose-dependently associated with lower LDL-c (P-trend 2.5 × 10-6 to 8.0 × 10-9), with lower risk of MI (P-trend 0.04-0.002) and lower risk of cardiovascular mortality (P-trend 0.005-0.004). These results were validated for LDL-c, risk of both coronary artery disease and MI in meta-analyses including from 194 000 to >700 000 participants. The variant is shown to increase CCDC93 protein stability, while overexpression of human CCDC93 decreases plasma LDL-c in mice. Conversely, CCDC93 ablation reduces LDL uptake as a result of reduced LDLR levels at the cell membrane., Conclusion: This study provides evidence that a common variant in CCDC93, encoding a protein involved in recycling of the LDLR, is associated with lower LDL-c levels, lower risk of MI and cardiovascular mortality., (© The Author(s) 2019. Published by Oxford University Press on behalf of the European Society of Cardiology.)
- Published
- 2020
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24. The hepatic WASH complex is required for efficient plasma LDL and HDL cholesterol clearance.
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Wijers M, Zanoni P, Liv N, Vos DY, Jäckstein MY, Smit M, Wilbrink S, Wolters JC, van der Veen YT, Huijkman N, Dekker D, Kloosterhuis N, van Dijk TH, Billadeau DD, Kuipers F, Klumperman J, von Eckardstein A, Kuivenhoven JA, and van de Sluis B
- Subjects
- Animals, Cholesterol, HDL blood, Cholesterol, LDL blood, Female, Liver chemistry, Male, Mice, Mice, Inbred C57BL, Mice, Transgenic, Scavenger Receptors, Class B metabolism, Cholesterol, HDL metabolism, Cholesterol, LDL metabolism, Liver metabolism, Microfilament Proteins genetics, Microfilament Proteins metabolism, Vesicular Transport Proteins genetics, Vesicular Transport Proteins metabolism
- Abstract
The evolutionary conserved Wiskott-Aldrich syndrome protein and SCAR homolog (WASH) complex is one of the crucial multiprotein complexes that facilitates endosomal recycling of transmembrane proteins. Defects in WASH components have been associated with inherited developmental and neurological disorders in humans. Here, we show that hepatic ablation of the WASH component Washc1 in chow-fed mice increases plasma concentrations of cholesterol in both LDLs and HDLs, without affecting hepatic cholesterol content, hepatic cholesterol synthesis, biliary cholesterol excretion, or hepatic bile acid metabolism. Elevated plasma LDL cholesterol was related to reduced hepatocytic surface levels of the LDL receptor (LDLR) and the LDLR-related protein LRP1. Hepatic WASH ablation also reduced the surface levels of scavenger receptor class B type I and, concomitantly, selective uptake of HDL cholesterol into the liver. Furthermore, we found that WASHC1 deficiency increases LDLR proteolysis by the inducible degrader of LDLR, but does not affect proprotein convertase subtilisin/kexin type 9-mediated LDLR degradation. Remarkably, however, loss of hepatic WASHC1 may sensitize LDLR for proprotein convertase subtilisin/kexin type 9-induced degradation. Altogether, these findings identify the WASH complex as a regulator of LDL as well as HDL metabolism and provide in vivo evidence for endosomal trafficking of scavenger receptor class B type I in hepatocytes.
- Published
- 2019
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25. Partial Deletion of Tie2 Affects Microvascular Endothelial Responses to Critical Illness in A Vascular Bed and Organ-Specific Way.
- Author
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Jongman RM, Zwiers PJ, van de Sluis B, van der Laan M, Moser J, Zijlstra JG, Dekker D, Huijkman N, Moorlag HE, Popa ER, Molema G, and van Meurs M
- Subjects
- Animals, E-Selectin genetics, E-Selectin metabolism, Endothelial Cells pathology, Inflammation chemically induced, Inflammation genetics, Inflammation metabolism, Inflammation pathology, Intercellular Adhesion Molecule-1 genetics, Intercellular Adhesion Molecule-1 metabolism, Leukocyte Common Antigens genetics, Leukocyte Common Antigens metabolism, Lipopolysaccharides toxicity, Mice, Mice, Knockout, Microvessels pathology, Organ Specificity, Receptor, TIE-2 genetics, Vascular Cell Adhesion Molecule-1 genetics, Vascular Cell Adhesion Molecule-1 metabolism, Endothelial Cells metabolism, Microvessels metabolism, Receptor, TIE-2 metabolism
- Abstract
Tyrosine kinase receptor (Tie2) is mainly expressed by endothelial cells. In animal models mimicking critical illness, Tie2 levels in organs are temporarily reduced. Functional consequences of these reduced Tie2 levels on microvascular endothelial behavior are unknown. We investigated the effect of partial deletion of Tie2 on the inflammatory status of endothelial cells in different organs. Newly generated heterozygous Tie2 knockout mice (exon 9 deletion, ΔE9/Tie2) exhibiting 50% reduction in Tie2 mRNA and protein, and wild-type littermate controls (Tie2), were subjected to hemorrhagic shock and resuscitation (HS + R), or challenged with i.p. lipopolysaccharide (LPS). Kidney, liver, lung, heart, brain, and intestine were analyzed for mRNA levels of adhesion molecules E-selectin, vascular cell adhesion molecule 1 (VCAM-1), and intercellular cell adhesion molecule 1 (ICAM-1), and CD45. Exposure to HS + R did not result in different expression responses of these molecules between organs from Tie2 or Tie2 mice and sham-operated mice. In contrast, the LPS-induced mRNA expression levels of E-selectin, VCAM-1, and ICAM-1, and CD45 in organs were attenuated in Tie2 mice when compared with Tie2 mice in kidney and liver, but not in the other organs studied. Furthermore, reduced expression of E-selectin and VCAM-1 protein, and reduced influx of CD45 cells upon LPS exposure, was visible in a microvascular bed-specific pattern in kidney and liver of Tie2 mice compared with controls. In contrast to the hypothesis that a disbalance in the Ang/Tie2 system leads to increased microvascular inflammation, heterozygous deletion of Tie2 is associated with an organ-restricted, microvascular bed-specific attenuation of endothelial inflammatory response to LPS.
- Published
- 2019
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26. The COMMD Family Regulates Plasma LDL Levels and Attenuates Atherosclerosis Through Stabilizing the CCC Complex in Endosomal LDLR Trafficking.
- Author
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Fedoseienko A, Wijers M, Wolters JC, Dekker D, Smit M, Huijkman N, Kloosterhuis N, Klug H, Schepers A, Willems van Dijk K, Levels JHM, Billadeau DD, Hofker MH, van Deursen J, Westerterp M, Burstein E, Kuivenhoven JA, and van de Sluis B
- Subjects
- Adaptor Proteins, Signal Transducing genetics, Animals, Atherosclerosis metabolism, Carrier Proteins genetics, Carrier Proteins metabolism, Cell Line, Cholesterol analysis, Chromatography, High Pressure Liquid, Cytoskeletal Proteins genetics, Gene Deletion, Gene Expression, HEK293 Cells, Hep G2 Cells, Humans, Liver chemistry, Liver metabolism, Low Density Lipoprotein Receptor-Related Protein-1, Male, Mice, Mice, Knockout, Protein Transport, Triglycerides analysis, Tumor Suppressor Proteins metabolism, Adaptor Proteins, Signal Transducing metabolism, Atherosclerosis prevention & control, Cholesterol, LDL blood, Cytoskeletal Proteins metabolism, Endosomes metabolism, Receptors, LDL metabolism
- Abstract
Rationale: COMMD (copper metabolism MURR1 domain)-containing proteins are a part of the CCC (COMMD-CCDC22 [coiled-coil domain containing 22]-CCDC93 [coiled-coil domain containing 93]) complex facilitating endosomal trafficking of cell surface receptors. Hepatic COMMD1 inactivation decreases CCDC22 and CCDC93 protein levels, impairs the recycling of the LDLR (low-density lipoprotein receptor), and increases plasma low-density lipoprotein cholesterol levels in mice. However, whether any of the other COMMD members function similarly as COMMD1 and whether perturbation in the CCC complex promotes atherogenesis remain unclear., Objective: The main aim of this study is to unravel the contribution of evolutionarily conserved COMMD proteins to plasma lipoprotein levels and atherogenesis., Methods and Results: Using liver-specific Commd1 , Commd6 , or Commd9 knockout mice, we investigated the relation between the COMMD proteins in the regulation of plasma cholesterol levels. Combining biochemical and quantitative targeted proteomic approaches, we found that hepatic COMMD1, COMMD6, or COMMD9 deficiency resulted in massive reduction in the protein levels of all 10 COMMDs. This decrease in COMMD protein levels coincided with destabilizing of the core (CCDC22, CCDC93, and chromosome 16 open reading frame 62 [C16orf62]) of the CCC complex, reduced cell surface levels of LDLR and LRP1 (LDLR-related protein 1), followed by increased plasma low-density lipoprotein cholesterol levels. To assess the direct contribution of the CCC core in the regulation of plasma cholesterol levels, Ccdc22 was deleted in mouse livers via CRISPR/Cas9-mediated somatic gene editing. CCDC22 deficiency also destabilized the complete CCC complex and resulted in elevated plasma low-density lipoprotein cholesterol levels. Finally, we found that hepatic disruption of the CCC complex exacerbates dyslipidemia and atherosclerosis in ApoE3*Leiden mice., Conclusions: Collectively, these findings demonstrate a strong interrelationship between COMMD proteins and the core of the CCC complex in endosomal LDLR trafficking. Hepatic disruption of either of these CCC components causes hypercholesterolemia and exacerbates atherosclerosis. Our results indicate that not only COMMD1 but all other COMMDs and CCC components may be potential targets for modulating plasma lipid levels in humans., (© 2018 American Heart Association, Inc.)
- Published
- 2018
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27. NF-κB p65 serine 467 phosphorylation sensitizes mice to weight gain and TNFα-or diet-induced inflammation.
- Author
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Riedlinger T, Dommerholt MB, Wijshake T, Kruit JK, Huijkman N, Dekker D, Koster M, Kloosterhuis N, Koonen DPY, de Bruin A, Baker D, Hofker MH, van Deursen J, Jonker JW, Schmitz ML, and van de Sluis B
- Subjects
- Aging metabolism, Aging pathology, Amino Acid Substitution genetics, Animals, Gene Expression Regulation, Gene Knock-In Techniques, Humans, Inflammation genetics, Inflammation pathology, Insulin metabolism, Liver metabolism, Liver pathology, Mice, Obesity metabolism, Obesity pathology, Phosphorylation, Serine metabolism, Transcription Factor RelA metabolism, Tumor Necrosis Factor-alpha metabolism, Weight Gain genetics, Aging genetics, Inflammation metabolism, Obesity genetics, Transcription Factor RelA genetics
- Abstract
The NF-κB family of transcription factors is essential for an effective immune response, but also controls cell metabolism, proliferation and apoptosis. Its broad relevance and the high connectivity to diverse signaling pathways require a tight control of NF-κB activity. To investigate the control of NF-κB activity by phosphorylation of the NF-κB p65 subunit, we generated a knock-in mouse model in which serine 467 (the mouse homolog of human p65 serine 468) was replaced with a non-phosphorylatable alanine (S467A). This substitution caused reduced p65 protein synthesis and diminished TNFα-induced expression of a selected group of NF-κB-dependent genes. Intriguingly, high-fat fed S467A mice displayed increased locomotor activity and energy expenditure, which coincided with a reduced body weight gain. Although glucose metabolism or insulin sensitivity was not improved, diet-induced liver inflammation was diminished in S467A mice. Altogether, this study demonstrates that phosphorylation of p65 serine 467 augment NF-κB activity and exacerbates various deleterious effects of overnutrition in mice., (Copyright © 2017 Elsevier B.V. All rights reserved.)
- Published
- 2017
- Full Text
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28. CCC- and WASH-mediated endosomal sorting of LDLR is required for normal clearance of circulating LDL.
- Author
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Bartuzi P, Billadeau DD, Favier R, Rong S, Dekker D, Fedoseienko A, Fieten H, Wijers M, Levels JH, Huijkman N, Kloosterhuis N, van der Molen H, Brufau G, Groen AK, Elliott AM, Kuivenhoven JA, Plecko B, Grangl G, McGaughran J, Horton JD, Burstein E, Hofker MH, and van de Sluis B
- Subjects
- Adolescent, Adult, Animals, Animals, Genetically Modified, Child, Child, Preschool, Chromatography, Liquid, Dogs, Female, Fluorescent Antibody Technique, HEK293 Cells, Humans, Immunoprecipitation, Male, Mice, Mice, Knockout, Middle Aged, Mutation, Protein Transport genetics, Transcriptome, Young Adult, Adaptor Proteins, Signal Transducing genetics, Cholesterol, LDL metabolism, Endosomes metabolism, Hypercholesterolemia genetics, Liver metabolism, Microfilament Proteins genetics, Proteins genetics, Receptors, LDL metabolism, Triglycerides metabolism
- Abstract
The low-density lipoprotein receptor (LDLR) plays a pivotal role in clearing atherogenic circulating low-density lipoprotein (LDL) cholesterol. Here we show that the COMMD/CCDC22/CCDC93 (CCC) and the Wiskott-Aldrich syndrome protein and SCAR homologue (WASH) complexes are both crucial for endosomal sorting of LDLR and for its function. We find that patients with X-linked intellectual disability caused by mutations in CCDC22 are hypercholesterolaemic, and that COMMD1-deficient dogs and liver-specific Commd1 knockout mice have elevated plasma LDL cholesterol levels. Furthermore, Commd1 depletion results in mislocalization of LDLR, accompanied by decreased LDL uptake. Increased total plasma cholesterol levels are also seen in hepatic COMMD9-deficient mice. Inactivation of the CCC-associated WASH complex causes LDLR mislocalization, increased lysosomal degradation of LDLR and impaired LDL uptake. Furthermore, a mutation in the WASH component KIAA0196 (strumpellin) is associated with hypercholesterolaemia in humans. Altogether, this study provides valuable insights into the mechanisms regulating cholesterol homeostasis and LDLR trafficking.
- Published
- 2016
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29. IL-1β and TGFβ2 synergistically induce endothelial to mesenchymal transition in an NFκB-dependent manner.
- Author
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Maleszewska M, Moonen JR, Huijkman N, van de Sluis B, Krenning G, and Harmsen MC
- Subjects
- Active Transport, Cell Nucleus immunology, Animals, Cellular Microenvironment drug effects, Endothelial Cells pathology, Epithelial-Mesenchymal Transition drug effects, Fibrosis immunology, Fibrosis pathology, Human Umbilical Vein Endothelial Cells, Humans, Inflammation immunology, Inflammation pathology, Interleukin-1beta pharmacology, Male, Mice, Transforming Growth Factor beta1 immunology, Transforming Growth Factor beta1 pharmacology, Transforming Growth Factor beta2 pharmacology, Cell Nucleus immunology, Cellular Microenvironment immunology, Endothelial Cells immunology, Epithelial-Mesenchymal Transition immunology, Interleukin-1beta immunology, NF-kappa B immunology, Transforming Growth Factor beta2 immunology
- Abstract
Endothelial to mesenchymal transition (EndMT) contributes to fibrotic diseases. The main inducer of EndMT is TGFβ signaling. TGFβ2 is the dominant isoform in the physiological embryonic EndMT, but its role in the pathological EndMT in the context of inflammatory co-stimulation is not known. The aim of this study was to investigate TGFβ2-induced EndMT in the context of inflammatory IL-1β signaling. Co-stimulation with IL-1β and TGFβ2, but not TGFβ1, caused synergistic induction of EndMT. Also, TGFβ2 was the only TGFβ isoform that was progressively upregulated during EndMT. External IL-1β stimulation was dispensable once EndMT was induced. The inflammatory transcription factor NFκB was upregulated in an additive manner by IL-1β and TGFβ2 co-stimulation. Co-stimulation also led to the nuclear translocation of NFκB which was sustained over long-term treatment. Activation of NFκB was indispensable for the co-induction of EndMT. Our data suggest that the microenvironment at the verge between inflammation (IL-1β) and tissue remodeling (TGFβ2) can strongly promote the process of EndMT. Therefore our findings provide new insights into the mechanisms of pathological EndMT., (Copyright © 2012 Elsevier GmbH. All rights reserved.)
- Published
- 2013
- Full Text
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30. The liver X receptor (LXR) and its target gene ABCA1 are regulated upon low oxygen in human trophoblast cells: a reason for alterations in preeclampsia?
- Author
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Plösch T, Gellhaus A, van Straten EM, Wolf N, Huijkman NC, Schmidt M, Dunk CE, Kuipers F, and Winterhager E
- Subjects
- ATP-Binding Cassette Transporters genetics, Anticholesteremic Agents pharmacology, Cell Line, Tumor, Cholesterol metabolism, Female, Gene Expression Regulation, Developmental, Humans, Hydrocarbons, Fluorinated pharmacology, Immunoblotting, Immunohistochemistry, In Vitro Techniques, Liver X Receptors, Orphan Nuclear Receptors agonists, Orphan Nuclear Receptors genetics, Oxygen administration & dosage, Placenta cytology, Pre-Eclampsia pathology, Pregnancy, RNA chemistry, RNA genetics, Reverse Transcriptase Polymerase Chain Reaction, Statistics, Nonparametric, Sulfonamides pharmacology, Trophoblasts cytology, ATP-Binding Cassette Transporters metabolism, Hypoxia metabolism, Orphan Nuclear Receptors metabolism, Oxygen metabolism, Placenta metabolism, Pre-Eclampsia metabolism, Trophoblasts metabolism
- Abstract
Objectives: The Liver X receptors (LXR) alpha and beta and their target genes such as the ATP-binding cassette (ABC) transporters have been shown to be crucially involved in the regulation of cellular cholesterol homeostasis. The aim of this study was to characterize the role of LXR alpha/beta in the human placenta under normal physiological circumstances and in preeclampsia., Study Design: We investigated the expression pattern of the LXRs and their target genes in the human placenta during normal pregnancy and in preeclampsia. Placental explants and cell lines were studied under different oxygen levels and pharmacological LXR agonists., Main Outcome Measures: Gene expressions (Taqman PCR) and protein levels (Western Blot) were combined with immunohistochemistry to analyze the expression of LXR and its target genes., Results: In the human placenta, LXRA and LXRB expression increased during normal pregnancy. This was paralleled by the expression of their prototypical target genes, e.g., the cholesterol transporter ABCA1. Interestingly, early-onset preeclamptic placentae revealed a significant upregulation of ABCA1. Culture of JAr trophoblast cells and human first trimester placental explants under low oxygen lead to increased expression of LXRA and ABCA1 which was further enhanced by the LXR agonist T0901317., Conclusions: LXRA together with ABCA1 are specifically expressed in the human placenta and can be regulated by hypoxia. Deregulation of this system in early preeclampsia might be the result of placental hypoxia and hence might have consequences for maternal-fetal cholesterol transport., (Copyright © 2010 Elsevier Ltd. All rights reserved.)
- Published
- 2010
- Full Text
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31. Pharmacological activation of LXR in utero directly influences ABC transporter expression and function in mice but does not affect adult cholesterol metabolism.
- Author
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van Straten EM, Huijkman NC, Baller JF, Kuipers F, and Plösch T
- Subjects
- Animals, Animals, Newborn, Anticholesteremic Agents pharmacology, Diet, Atherogenic, Embryo, Mammalian, Female, Fetal Development drug effects, Fetal Development genetics, Gene Expression Regulation, Developmental drug effects, Lipid Metabolism drug effects, Lipid Metabolism genetics, Liver X Receptors, Male, Mice, Mice, Inbred C57BL, Mice, Transgenic, Orphan Nuclear Receptors, Pregnancy, Prenatal Exposure Delayed Effects genetics, ATP-Binding Cassette Transporters genetics, ATP-Binding Cassette Transporters physiology, Cholesterol metabolism, DNA-Binding Proteins agonists, Hydrocarbons, Fluorinated pharmacology, Prenatal Exposure Delayed Effects metabolism, Receptors, Cytoplasmic and Nuclear agonists, Sulfonamides pharmacology
- Abstract
Cholesterol is critical for several cellular functions and essential for normal fetal development. Therefore, its metabolism is tightly controlled during all life stages. The liver X receptors-alpha (LXRalpha; NR1H3) and -beta (LXRbeta; NR1H2) are nuclear receptors that are of key relevance in coordinating cholesterol and fatty acid metabolism. The aim of this study was to elucidate whether fetal cholesterol metabolism can be influenced in utero via pharmacological activation of LXR and whether this would have long-term effects on cholesterol homeostasis. Administration of the LXR agonist T0901317 to pregnant mice via their diet (0.015% wt/wt) led to induced fetal hepatic expression levels of the cholesterol transporter genes Abcg5/g8 and Abca1, higher plasma cholesterol levels, and lower hepatic cholesterol levels compared with controls. These profound changes during fetal development did not affect cholesterol metabolism in adulthood nor did they influence coping with a high-fat/high-cholesterol diet. This study shows that the LXR system is functional in fetal mice and susceptible to pharmacological activation. Despite massive changes in fetal cholesterol metabolism, regulatory mechanisms involved in cholesterol metabolism return to a "normal" state in offspring and allow coping with a high-fat/high-cholesterol diet.
- Published
- 2008
- Full Text
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32. A stable full-length yellow fever virus cDNA clone and the role of conserved RNA elements in flavivirus replication.
- Author
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Bredenbeek PJ, Kooi EA, Lindenbach B, Huijkman N, Rice CM, and Spaan WJM
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- 3' Untranslated Regions chemistry, 3' Untranslated Regions genetics, Animals, Cell Line, Conserved Sequence, Cricetinae, Escherichia coli genetics, RNA, Viral biosynthesis, RNA, Viral chemistry, RNA, Viral genetics, Transcription, Genetic, Transfection, Viral Plaque Assay, Yellow fever virus genetics, Yellow fever virus physiology, Cloning, Molecular, DNA, Complementary, Virus Replication, Yellow fever virus pathogenicity
- Abstract
Yellow fever virus (YF) is the prototype member of the Flavivirus genus. Here, we report the successful construction of a full-length infectious cDNA clone of the vaccine strain YF-17D. YF cDNA was cloned into a low-copy-number plasmid backbone and stably maintained in several E. coli strains. Transcribed RNAs had a specific infectivity of 10(5)-10(6) p.f.u. ( micro g RNA)(-1), and the resulting virus exhibited growth kinetics, plaque morphology and proteolytic processing similar to the parental virus in cell culture. This clone was used to analyse the importance of conserved flavivirus RNA sequences and the 3' stem-loop structure in virus replication. The conserved sequences 5'CS and CS1, as well as the 3' stem-loop structure, were found to be essential for virus replication in cell culture, whereas the conserved sequence CS2 and the region containing YF-specific repeated sequences were dispensable. This infectious clone will aid future studies on YF replication and pathogenesis, as well as facilitate the development of YF-17D-based recombinant vaccines.
- Published
- 2003
- Full Text
- View/download PDF
33. Genomics of the human carnitine acyltransferase genes.
- Author
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van der Leij FR, Huijkman NC, Boomsma C, Kuipers JR, and Bartelds B
- Subjects
- Carnitine O-Acetyltransferase genetics, Carnitine O-Palmitoyltransferase genetics, Chromosome Mapping, DNA, Complementary genetics, Exons, Genome, Human, Humans, Introns, Isoenzymes genetics, Phylogeny, Promoter Regions, Genetic, Pseudogenes, Carnitine Acyltransferases genetics
- Abstract
Five genes in the human genome are known to encode different active forms of related carnitine acyltransferases: CPT1A for liver-type carnitine palmitoyltransferase I, CPT1B for muscle-type carnitine palmitoyltransferase I, CPT2 for carnitine palmitoyltransferase II, CROT for carnitine octanoyltransferase, and CRAT for carnitine acetyltransferase. Only from two of these genes (CPT1B and CPT2) have full genomic structures been described. Data from the human genome sequencing efforts now reveal drafts of the genomic structure of CPT1A and CRAT, the latter not being known from any other mammal. Furthermore, cDNA sequences of human CROT were obtained recently, and database analysis revealed a completed bacterial artificial chromosome sequence that contains the entire CROT gene and several exons of the flanking genes P53TG and PGY3. The genomic location of CROT is at chromosome 7q21.1. There is a putative CPT1-like pseudogene in the carnitine/choline acyltransferase family at chromosome 19. Here we give a brief overview of the functional relations between the different carnitine acyltransferases and some of the common features of their genes. We will highlight the phylogenetics of the human carnitine acyltransferase genes in relation to the fungal genes YAT1 and CAT2, which encode cytosolic and mitochondrial/peroxisomal carnitine acetyltransferases, respectively., (Copyright 2000 Academic Press.)
- Published
- 2000
- Full Text
- View/download PDF
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